Elevated PGE2 Research Articles

Scutellaria baicalensis Alleviates Cantharidin-Induced Rat Hemorrhagic Cystitis through Inhibition of Cyclooxygenase-2 Overexpression

Cantharidin, an active component in mylabris, is used in traditional Chinese medicine (TCM) to treat scabies and hepatoma, but accompanied by hemorrhagic cystitis. Evidence shows that cantharidin induces human bladder carcinoma cell death through COX-2 overexpression in vitro. In TCM, Scutellaria baicalensis is usually used to cure mylabris-induced hematuria. This work was undertaken to determine the mechanisms of cantharidin-induced rat hemorrhagic cystitis and explore the uroprotective effect of S. baicalensis. In vitro results showed cantharidin could induce cytotoxicity through prostaglandin (PG)E2 overproduction of T24 cells. Boiling-water extract of S. baicalensis (SB-WE) could significantly inhibit PGE2 production and COX-2 expression in lipo-polysaccharide-induced RAW 264.7 cells, indicating obvious anti-inflammatory abilities. In vivo results indicated that cantharidin caused rat hemorrhagic cystitis with hematuria via c-Fos and COX-2 overexpression. SB-WE was given orally to cantharidin-treated rats, whereby hematuria level, elevated PGE2 and COX-2 protein overexpression were significantly and dose-dependently inhibited by SB-WE. The anti-inflammatory components of SB-WE are baicalin and wogonin, whose contents were 200.95 ± 2.00 and 31.93 ± 0.26 μg/mg, respectively. In conclusion, cantharidin induces rat cystitis through c-Fos and COX-2 over-expression and S. baicalensis can prevent the resulting hematuria because of its anti-inflammatory effects.

Abstract 543: Estrogen metabolism within prostatic micro-milieu: insights to prostatic carcinogenesis

Abstract Prostatic cancer is amongst the most common neoplasm in men. Inflammation and accelerated proliferation are important underlying causes for prostatic carcinogenesis. The role of estrogen in prostate physiology and pathophysiology has been growing rapidly. However some estrogen metabolites are more potent relative to parent estrogen. In the current work, the reciprocal role between inflammation and estrogen metabolism in prostatic microenvironment was investigated. Prostatitis was induced in normal and estrogen treated male rats by intraprostatic injection of 2% carragenan solution. After 48 h, animals were euthanized and prostate glands were collected. Prostatic inflammation and proliferation were confirmed by gross visual, histological and elevated TNF-α, PGE2 and cyclin-D1. Expression of estrogen metabolizing enzymes was assessed using real time PCR and estrogen metabolites within prostate tissue were assayed using LC-MS/MS. Inflammation combined with estrogen exposure showed exaggerated inflammatory and proliferative response. Estrogen metabolizing enzymes were over expressed in response to prostatic inflammation with sequential accumulation of catechol estrogens within prostatic tissues. Catechol estrogens were found to accelerate prostatic epithelial cell proliferation and alter their genetic stability. In conclusion, the current work provides experimental evidence that prostatic inflammation enhances estrogen metabolism to generate catechol estrogens within prostatic microenvironment. These may contribute in prostatic inflammation and accelerated proliferation and subsequently prostatic carcinogenesis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 543. doi:1538-7445.AM2012-543

Potently Immunosuppressive 5-Fluorouracil–Resistant Mesenchymal Stromal Cells Completely Remit an Experimental Autoimmune Disease

We treated mice with 5-fluorouracil (5-FU) to isolate a quiescent and undifferentiated mesenchymal stromal cell (MSC) population from the bone marrow. We examined these 5-FU-resistant MSCs (5-FU-MSCs) free from hematopoietic components for CFU fibroblasts (CFU-Fs) and assessed their immunosuppressive potential in vitro and in vivo. We differentiated fibroblastic CFU-Fs (Fibro-CFU-Fs) from mixed CFU-Fs, based on the absence of in situ expression of CD11b and CD45 hematopoietic markers, as well as on their differentiation capacity. Fibro-CFU-Fs were associated with increased numbers of large-sized Fibro-CFU-Fs (≥9 mm(2)) that displayed enhanced capacity for differentiation into adipogenic and osteogenic mesenchymal lineages. Administration of these 5-FU-resistant CD11b(-)CD45(-) MSCs 6 d after myelin oligodendrocyte glycoprotein (MOG) immunization completely remitted MOG-induced experimental autoimmune encephalomyelitis after initial development of mild disease. The remission was accompanied by reduced CNS cellular infiltration and demyelination, as well as a significant reduction in anti-MOG Ab and splenocyte proliferation to MOG. MOG-stimulated splenocytes from these mice showed elevated levels of Th2 cytokines (IL-4, IL-5, and IL-6) and decreased IL-17. Compared with untreated MSCs, 5-FU-MSCs demonstrated potent immunosuppression of Con A-stimulated splenocytes in vitro, even at a 1:320 MSC/splenocyte ratio. Immunosuppression was accompanied by elevated IL-1ra, IL-10, and PGE(2). Blocking IL-1ra, IL-10, and PGE(2), but not IL-6, heme oxygenase-1, and NO, attenuated 5-FU-MSC-induced immunosuppression. Together, our findings suggested that immunosuppression by 5-FU-MSC is mediated by a combination of elevated IL-1ra, IL-10, and PGE(2), anti-inflammatory Th2 cytokines, and decreased IL-17. Our findings suggested that 5-FU treatment identifies a population of potently immunosuppressive 5-FU-MSCs that have the potential to be exploited to remit autoimmune diseases.

Thirst deficits in aged rats are reversed by dietary omega-3 fatty acid supplementation

During heat waves many elderly individuals die as a consequence of dehydration. This is partially due to deficits in mechanisms controlling thirst. Reduced thirst following dipsogenic stimuli is well documented in aged humans and rodents. Low in vivo long-chain omega-3 fatty acid levels, as can occur in aging, have been shown to alter body fluid and sodium homeostasis. Therefore, the effect of dietary omega-3 fatty acid supplementation on drinking responses in aged rats was examined. Omega-3 fatty acids reversed thirst deficits in aged rats following dehydration and hypertonic stimuli; angiotensin (ANG) II induced drinking was unaffected in aged rats. Plasma atrial natriuretic peptide (ANP) and arginine vasopressin (AVP) were altered with age, but not affected by diet. Aged omega-3 fatty acid deficient animals displayed increased hypothalamic cytosolic phospholipase A2 (cPLA2), cyclooxygenase (COX)-2, and prostaglandin E (PGE) synthase messenger (m)RNA expression, compared with animals that received omega-3 fatty acids. The aged low omega-3 fatty acid fed animals had significantly elevated hypothalamic PGE2 compared with all other groups. Hypothalamic PGE2 was negatively correlated with drinking induced by both dehydration and hypertonicity. The results indicate that PGE2 may be the underlying mechanism of the reduced thirst observed in aging.

Elevated Plasma Oxidized Lipids in Severe Pulmonary Hypertension are Fully Restored by Estrogen Therapy

Recently we reported that estrogen (E2) rescues pre-existing severe pulmonary hypertension (PH). PH is associated with pulmonary vascular remodelling. Vascular disorders result from complex interactions between oxidized-lipoproteins, monocytes/macrophages, injured endothelium and smooth muscle cells. Biological oxidation products of arachidonic acid, including prostaglandins (PGs), thromboxanes (TXs), hydroxyeicosatetraenoic acids (HETEs) and hydroxyoctadecadienoic acids (HODEs) play an important role in the pathogenesis of atherosclerosis, yet their role in PH is not fully known. Here we investigated whether PH is associated with increased plasma levels of oxidized-lipids, and if E2 can restore their levels. To induce PH, rats received monocrotaline (MCT, 60mg/kg, s.c.). Severe PH was established 21-days after MCT (PH, n=8). E2 (42.5ug/kg/day, s.c., n=6) was given from day-21 to 30. Saline treated rats served as control (CTRL, n=7). Cardiac catheterization was performed terminally to record RV-pressure (RVP). Plasma HODEs, HETEs, TXs and PGs were determined by mass-spectrometry. p<0.05 was considered significant. Values were mean±SE. Rats developed severe PH [RVP=68±2 vs. 31±2 mmHg in CTRL, p<0.01). E2 rescued PH (RVP=38±1 mmHg, p<0.01 vs. PH). PH was associated with increased plasma levels (ng/ml of plasma) of 9-HODE (2217±381 vs. 949±255 in CTRL, p<0.05) and 13-HODE (4347±439 vs. 1871±609 in CTRL, p<0.05). E2 reversed PH-induced increase in HODEs (9-HODE=976±259; 13-HODE=2469±531; all p<0.05 vs. PH). PH led to significantly elevated plasma 5-HETE (938±77 vs. 663±15), 12-HETE (9720±972 vs. 6633±522) and 15-HETE (818±62 vs. 534±7; all p<0.05 vs. CTRL). E2 fully reversed the increase in HETEs (5-HETE=708±37; 12-HETE=6981±248; 15-HETE=622±50; p<0.05 vs. PH). PH also led to elevated plasma TXB2 (3.5±0.2 vs. 2.8±0.1), PGD2 (98±11 vs. 35±7) and PGE2 (27±1 vs. 13±2; all p<0.05 vs. CTRL) and E2 restored their levels (TXB2=2.6±0.2; PGD2=54±10; PGE2=19±2; all p<0.05). In conclusion, severe PH is associated with increased plasma levels of oxidized lipids that are fully restored by E2.

Exogenous ghrelin attenuates endotoxin fever in rats

Ghrelin is a gut-derived peptide that plays a role in energy homeostasis. Recent studies have implicated ghrelin in systemic inflammation, showing increased plasma ghrelin levels after endotoxin (lipopolysaccharide, LPS) administration. The aims of this study were (1) to test the hypothesis that ghrelin administration affects LPS-induced fever; and (2) to assess the putative effects of ghrelin on plasma corticosterone secretion and preoptic region prostaglandin (PG) E2 levels in euthermic and febrile rats. Rats were implanted with a temperature datalogger capsule in the peritoneal cavity to record body core temperature. One week later, they were challenged with LPS (50μg/kg, intraperitoneal, i.p.) alone or combined with ghrelin (0.1mg/kg, i.p.). In another group of rats, plasma corticosterone and preoptic region PGE2 levels were measured 2h after injections. In euthermic animals, systemic administration of ghrelin failed to elicit any thermoregulatory effect, and caused no significant changes in basal plasma corticosterone and preoptic region PGE2 levels. LPS caused a typical febrile response, accompanied by increased plasma corticosterone and preoptic PGE2 levels. When LPS administration was combined with ghrelin fever was attenuated, corticosterone secretion further increased, and the elevated preoptic PGE2 levels were relatively reduced, but a correlation between these two variables (corticosterone and PGE2) failed to exist. The present data add ghrelin to the neurochemical milieu controlling the immune/thermoregulatory system acting as an antipyretic molecule. Moreover, our findings also support the notion that ghrelin attenuates fever by means of a direct effect of the peptide reducing PGE2 production in the preoptic region.

Acute Tumor Lysis Syndrome. Modulation of Cyclooxygenase-2, Alpha V Beta Integrin and Microsomal Prostaglandin E Synthase-1 In Human Immunodeficiency Virus-1 (HIV-1) Squamous Cell Carcinoma (SCC) of the Penis with Selenomethionine/Nsaids

AbstractAbstract 4880Invasive squamous cell carcinoma of the penis overexpress Cyclooxygenase-2 (cox-2), Microsomal Prostaglandin E Synthase-1 and Alpha V Beta Integrins. Trace minerals selenium (selenomethionine) is deficient in many neoplasia conditions especially SCC of the penis in HIV-1 patients. Selenium, an essential trace element, has been shown to inhibit tumorigenesis. Hence, selenium and arachidonic acid metabolism may be linked to SCC. Penile cancer (SCC histology) displays elevated COX-2 and PGE2 can be regulated by selenium supplementation.We present a 47 We present a 47 yo male with HIV-1, CD4-133/cumm, VL of 95,000 copies/ml with T4 advanced stage SCC of the penis post XRT, surgical skin graft with extensive lymphedema, pain and KPS score of 40–50%. CT scan of Abdomen/Pelvis was c/w extensive disease of his right inguinal region, bladder compressed and displaced into left pelvis and tissue massof the right and left groin with ill defined nodular densities within RLL and left hepatic lobe. Bone marrow was hypocellular, M:E ratio of 8:1 with foamy macrophages. When treated for selenium deficient level and pain with selenomethionine and COX-2 inhibitor, acute tumor lysis syndrome developed with hyperkalemia, hyperuricemia and hypocalcemia defervesing with aggressive therapy with hydration, alkalinization and allopurinol. Conclusion:Overexpression of COX-2, mPEGS-1, alpha V beta integrin leads to phenotypic changes in epithelial cells that could enhance tumorigenesis. Selenium supplementation and COX-2 inhibition may help regulate COX-2 alternating cellular adhesion and promote apoptosis in SCC of the penis and upon treatment of these HIV-1 patients, with heavy tumor burden, tumor lysis syndrome along with disease Invasive squamous cell carcinoma of the penis overexpress Cyclooxygenase-2 (cox-2), Microsomal Prostaglandin E Synthase-1 and Alpha V Beta Integrins. Trace minerals selenium (selenomethionine) is deficient in many neoplasia conditions and PGE2 can be regulated by selenium supplementation. especially SCC of the penis in HIV-1 patients. Selenium, an essential trace element, has been shown to inhibit tumorigenesis. Hence, selenium and arachidonic acid metabolism may be linked to SCC. Penile cancer (SCC histology) displays elevated COX-2. Disclosures:No relevant conflicts of interest to declare.

Protective actions of des‐aspartate‐angiotensin I in mice model of CEES‐induced lung intoxication

The present study investigated the protective actions of des-aspartate-angiotensin I (DAA-I) in mice that were intranasally administered 2-chloroethyl ethyl sulfide (CEES), a half sulfur mustard. The protection was dose-dependent, and an oral dose of 75 mg kg⁻¹ per day administered 18 h post exposure and for the following 13 days, offered maximum protection that increased survival by a third. DAA-I attenuated the early processes of inflammation seen in the CEES-inoculated mice. DAA-I attenuated (i) elevated pulmonary ROS, and gp91-phox protein of NADPH oxidase, a non phagocytic enzyme that generates superoxide and subsequent ROS; (ii) intercellular adhesion molecule-1 (ICAM⁻¹) that is involved in the extravasation of circulating leucocytes; and (iii) myeloperoxidase activity, which is a surrogate enzymatic measurement of neutrophil infiltration. These actions led to improved histological lung structures, and survival of type-1 pneumocytes. The action of DAA-I on animal survival was blocked by losartan, a selective angiotensin AT1 receptor blocker, indicting that the AT1 receptor mediates the protection. The presence of elevated PGE2 and PGI2 in lung supernatants of DAA-I treated CEES-inoculated mice indicates that the two prostaglandins are involved in signaling the protective actions of DAA-I. This finding complements earlier studies showing that DAA-I acts on an indomethacin-sensitive angiotensin AT1 receptor. The findings of the present study are the first demonstration of an angiotensin peptide as an effective antidote for CEES intoxication. DAA-I is also an effective therapeutic intervention against CEES that was instituted at 18 h post exposure, and challenges conventional assumptions of limited efficacy with delayed action against alkylating agents.

Proinflammatory-Activated Trigeminal Satellite Cells Promote Neuronal Sensitization: Relevance for Migraine Pathology

BackgroundMigraine is a complex, chronic, painful, neurovascular disorder characterized by episodic activation of the trigeminal system. Increased levels of calcitonin gene-related peptide (CGRP) are found at different levels during migraine attacks. Interestingly, CGRP is also released within the trigeminal ganglia suggesting possible local effects on satellite cells, a specialized type of glia that ensheaths trigeminal neurons. CGRP was shown to enhance satellite-cell production of interleukin 1β (IL-1β), while trigeminal neurons express an activity-dependent production of nitric oxide (NO). Thus, in the present study we tested the hypothesis that IL-1β and NO induce trigeminal satellite cell activation, and that once activated these cells can influence neuronal responses.ResultsPrimary cultures of rat trigeminal satellite cells isolated from neuronal cultures were characterized in vitro. Cyclooxygenase (COX) expression and activity were taken as a marker of glial pro-inflammatory activation. Most of the experiments were carried out to characterize satellite cell responses to the two different pro-inflammatory stimuli. Subsequently, medium harvested from activated satellite cells was used to test possible modulatory effects of glial factors on trigeminal neuronal activity. IL-1β and the NO donor diethylenetriamine/nitric oxide (DETA/NO) elevated PGE2 release by satellite cells. The stimulatory effect of IL-1β was mediated mainly by upregulation of the inducible form of COX enzyme (COX2), while NO increased the constitutive COX activity. Regardless of the activator used, it is relevant that short exposures of trigeminal satellite cells to both activators induced modifications within the cells which led to significant PGE2 production after removal of the pro-inflammatory stimuli. This effect allowed us to harvest medium from activated satellite cells (so-called 'conditioned medium') that did not contain any stimulus, and thus test the effects of glial factors on neuronal activation. Conditioned medium from satellite cells activated by either IL-1β or NO augmented the evoked release of CGRP by trigeminal neurons.ConclusionThese findings indicate that satellite cells contribute to migraine-related neurochemical events and are induced to do so by autocrine/paracrine stimuli (such as IL-1β and NO). The responsiveness of IL-1β to CGRP creates the potential for a positive feedback loop and, thus, a plurality of targets for therapeutic intervention in migraine.

Celecoxib has Potent Antitumour Effects as a Single Agent and in Combination with BCG Immunotherapy in a Model of Urothelial Cell Carcinoma

ObjectivesProstaglandin E2 (PGE2) is a potent immune modulator and known to suppress both tumour antigen-specific helper T (TH1) cells and the generation of cytotoxic T lymphocytes (CTLs). We hypothesised that a combination of the cyclooxygenase 2 (COX-2) selective inhibitor celecoxib and intravesical bacillus Calmette-Guérin (BCG), an effective tumour immunoprophylaxis and ablative therapy for non–muscle-invasive bladder cancer, would be more effective than BCG alone. MethodsWe assessed urinary levels of PGE2 in humans receiving BCG and in a murine model of urothelial cell carcinoma (UCC). The cytokine response to BCG plus celecoxib was assessed in murine dendritic cells (DCs) in vitro and tumour ablation was assessed in an orthotopic MBT2 murine bladder cancer model. ResultsAdministration of intravesical BCG resulted in elevated urinary PGE2 levels in patients with high-grade superficial UCC and in a mouse model of UCC. In vitro, activation of DCs with BCG stimulated COX-2 up-regulation and release of the archetypal tolerogenic factors, PGE2 and interleukin 10. In a superficial mouse model of UCC, combination of celecoxib and intravesical BCG therapy resulted in increased tumour infiltration of CD4+ T cells and improved efficacy when compared to BCG alone. Further, celecoxib demonstrated marked antitumour efficacy when administered in the absence of BCG immunotherapy. ConclusionsThis study demonstrates that a combination strategy involving BCG immunotherapy and celecoxib may be more therapeutically beneficial than stand-alone intravesical therapy.

Letter to the editor

Hambek et al1 demonstrated that serum PGE2 levels were inversely correlated with T classification in patients with squamous cell carcinoma of the head and neck. Elevated serum PGE2 levels were primarily seen in patients with T1 tumors. Although they report a significant difference between serum PGE2 levels in patients with T2/3 and T4 tumors, this seems improbable given the large overlap of the box and whisker plots (Figure 3). They hypothesized that this inverse correlation was due to tumor-derived PGE2 associated with the process of neovascularization during the proliferative phase of tumor growth. In our studies of cultured squamous cell carcinoma cell lines, PGE2 production by tumor cells was not associated with tumor site, stage, or 2-year disease-free survival.2 In studies of tumor-infiltrating mononuclear cells (TIMC), advanced N stage (but not T stage) was associated with decreased PGE2 production by TIMC.3 Although we did not observe an association of tumor tissue levels of PGE2 with tumor stage, increased levels of PGE2 were associated with increased 2-year disease-free survival.4 Collectively, these findings also suggest that increased production of PGE2 is a favorable prognostic sign and may be associated with earlier stage disease. In their article, Hambek et al do not establish the source of the PGE2 in the serum. Our own observations suggest that tumor cells are not the major source of PGE2; rather, it is derived from tumor-infiltrating and circulating mononuclear cells.2-4 An alternative explanation for their observations is that an inflammatory response to tumors in the early stage of tumor growth, perhaps induced by tumor-derived factors, results in increased production of PGE2 by inflammatory cells. As tumors progress, there is suppression of the host inflammatory response resulting in less PGE2 production. The results of cell culture analyses (Figure 1) in their study may not reflect the complex environment of tumor–host interactions in vivo. Subsequent studies in vivo using murine models of squamous cell carcinoma also support the hypothesis that PGE2 is derived from a host inflammatory response to the tumor and may be beneficial. Inhibition of tumor growth in mice treated with a cyclo-oxygenase inhibitor was associated with increased tumor tissue levels of PGE2 and lymphocytic infiltration.5 Peritumoral injections of tumors with PGE2 significantly inhibited tumor growth.6 Hambek et al also observed normalization of serum PGE2 levels in follow-up and suggest that serum PGE2 levels may be useful as a tumor marker. Do they have data comparing patients who developed recurrent disease or a second primary in follow-up? If serum PGE2 is derived from mononuclear cells rather than tumor cells, the utility of serum PGE2 as a tumor marker may be limited depending on the status of the immune system in patients who develop recurrent disease. I suspect that persistent immunosuppression will prevent significant PGE2 production by mononuclear cells and serum PGE2 levels will remain low.

PUFA and flavonoid actions on PGE 2 production in human epidermal keratinocytes

The purpose of this research is to determine if n-3 PUFA and flavonoids (catechins and quercetin) reduce biomarkers of inflammation in human epidermal keratinocytes. The current goal examined how anti-inflammatory n-3 and pro-inflammatory n-6 PUFA altered fatty acid composition and PGE2 production in cultures of epidermal keratinocytes HEK001 and HaCaT. In expt I, HEK001 and HaCaT cells were treated with three levels (5, 10, and 25 μM) of individual PUFA (LA, LNA, AA, EPA, or DHA). In expt II, both cell lines were treated with combinations of AA:EPA and AA:DHA at ratios of 3:1, 2:1, 1:1, 1:2 and 1:3 at a constant concentration of 12 μM. In expt III, the effects of PUFA enrichment on PGE2 production and the uptake of tea catechins were evaluated in HEK001 cells. Epigallocatechin gallate was found in cells administered with 10 μM of catechins after 3 hr. Analysis showed that these cells were extremely low in essential fatty acids. Enrichment with linoleic or linolenic acids did not significantly increase their respective 20 C PUFA. All 20 C PUFA treatments resulted in a dramatic and consistent elevation in the PUFA used for the enrichment following a graded response. AA treatment consistently elevated PGE2 production in HEK001 cells. The PUFA levels used here are physiological and should be tested in vivo.

APRIL and BAFF Promote Increased Viability of Replicating Human B2 Cells via Mechanism Involving Cyclooxygenase 2

Of relevance to both protective and pathogenic responses to Ag is the recent finding that soluble molecules of the innate immune system, i.e., IL-4, B cell-activation factor of the TNF family (BAFF), and C3, exhibit significant synergy in promoting the clonal expansion of human B2 cells following low-level BCR ligation. Although IL-4, BAFF, and C3dg each contribute to early cell cycle entry and progression to S phase, only BAFF promotes later sustained viability of progeny needed for continued cycling. The present study sought to further clarify the mechanisms for BAFF's multiple functions. By comparing BAFF and a proliferation-inducing ligand (APRIL) efficacy at different stages in the response (only BAFF binds BR3; both bind transmembrane activator and calcium modulator and cyclophilin ligand interactor (TACI) and B cell maturation Ag, the early role was attributed to BR3, while the later role was attributed to TACI/B cell maturation Ag. Importantly, BAFF- and APRIL-promoted viability of cycling lymphoblasts was associated with sustained expression of cyclooxygenase 2 (COX-2), the rate-limiting enzyme for PGE2 synthesis, within replicating cells. Supernatants of cultures with BAFF and APRIL contained elevated PGE2. Although COX-2 inhibitors diminished daughter cell viability, exogenous PGE2 (1-1000 nM) increased the viability and recovery of lymphoblasts. Increased yield of viable progeny was associated with elevated Mcl-1, suggesting that a BAFF/APRIL --> TACI --> COX-2 --> PGE2--> Mcl-1 pathway reduces activation-related, mitochondrial apoptosis in replicating human B2 cell clones.

Single Periocular Injection of Celecoxib-PLGA Microparticles Inhibits Diabetes-Induced Elevations in Retinal PGE2, VEGF, and Vascular Leakage

To determine whether celecoxib inhibits VEGF secretion from ARPE-19 cells and to investigate further the safety and effectiveness of periocular celecoxib-poly (lactide-co-glycolide; PLGA) microparticles in inhibiting elevations in retinal PGE(2), VEGF, and blood-tissue barrier leakage at the end of 60 days in a streptozotocin diabetic rat model. VEGF mRNA and protein expression in ARPE-19 cells was evaluated in the presence of 0 to 10 microM celecoxib, and cytotoxicity of celecoxib on ARPE-19 and RF6A cells was evaluated over a 0- to 100-microM concentration range. Celecoxib-PLGA microparticles were prepared by a modified solvent evaporation technique, sterilized by 25 kGy of gamma-irradiation, and characterized for size, zeta potential, drug loading, and in vitro release. Normal and streptozotocin-diabetic male Sprague-Dawley rats were divided into five groups: normal, diabetic, diabetic+placebo, normal+celecoxib, and diabetic+celecoxib. Phosphate-buffered saline (PBS) containing celecoxib-PLGA microparticles, placebo PLGA microparticles, or plain PBS in one eye was injected into the posterior subconjunctival (periocular) space in rats under anesthesia. Sixty days after administration, the animals were killed, and retinal PGE2 secretion, VEGF protein, and blood-retinal barrier leakage were estimated. Blood cell counts, blood chemistry and histology were used to assess the safety of the microparticulate system. Celecoxib (up to 25 microM) did not cause significant cytotoxicity in ARPE-19 or RF6A cells. Nanomolar concentrations of celecoxib reduced VEGF mRNA and VEGF protein secretion. Celecoxib-PLGA microparticles (diameter: 1140 +/- 15 nm), containing 14.93% +/- 0.21% of celecoxib sustained in vitro drug release and in vivo drug levels in the retina for 60 days. Diabetes elevated PGE2 secretion, VEGF protein, the vitreous-plasma protein ratio, and blood-retinal barrier leakage by 3-, 1.7-, 3.1-, and 2.7-fold, and celecoxib-PLGA microparticles significantly reduced these elevations by 40%, 50%, 40%, and 50%, respectively. Neither the placebo-treated eyes nor the contralateral eyes in celecoxib-PLGA microparticle-treated rats showed significant effects. Celecoxib-PLGA or placebo-PLGA particles had no effect on the body weight or blood sugar level of rats. The celecoxib-PLGA microparticles did not cause any changes in blood cell counts or chemistry and caused no histopathological damage to the retina or periocular tissues. Nanomolar concentrations of celecoxib can inhibit VEGF mRNA and protein expression from ARPE-19 cells. Periocular celecoxib microparticles are useful sustained drug delivery systems for inhibiting diabetes-induced elevations in PGE2, VEGF, and blood-retinal barrier leakage. The periocular celecoxib-PLGA microparticles are safe and do not cause any damage to the retina.

Hyperresponsiveness to inhaled but not intravenous methacholine during acute respiratory syncytial virus infection in mice

BackgroundTo characterise the acute physiological and inflammatory changes induced by low-dose RSV infection in mice.MethodsBALB/c mice were infected as adults (8 wk) or weanlings (3 wk) with 1 × 105 pfu of RSV A2 or vehicle (intranasal, 30 μl). Inflammation, cytokines and inflammatory markers in bronchoalveolar lavage fluid (BALF) and airway and tissue responses to inhaled methacholine (MCh; 0.001 – 30 mg/ml) were measured 5, 7, 10 and 21 days post infection. Responsiveness to iv MCh (6 – 96 μg/min/kg) in vivo and to electrical field stimulation (EFS) and MCh in vitro were measured at 7 d. Epithelial permeability was measured by Evans Blue dye leakage into BALF at 7 d. Respiratory mechanics were measured using low frequency forced oscillation in tracheostomised and ventilated (450 bpm, flexiVent) mice. Low frequency impedance spectra were calculated (0.5 – 20 Hz) and a model, consisting of an airway compartment [airway resistance (Raw) and inertance (Iaw)] and a constant-phase tissue compartment [coefficients of tissue damping (G) and elastance (H)] was fitted to the data.ResultsInflammation in adult mouse BALF peaked at 7 d (RSV 15.6 (4.7 SE) vs. control 3.7 (0.7) × 104 cells/ml; p < 0.001), resolving by 21 d, with no increase in weanlings at any timepoint. RSV-infected mice were hyperresponsive to aerosolised MCh at 5 and 7 d (PC200 Raw adults: RSV 0.02 (0.005) vs. control 1.1 (0.41) mg/ml; p = 0.003) (PC200 Raw weanlings: RSV 0.19 (0.12) vs. control 10.2 (6.0) mg/ml MCh; p = 0.001). Increased responsiveness to aerosolised MCh was matched by elevated levels of cysLT at 5 d and elevated VEGF and PGE2 at 7 d in BALF from both adult and weanling mice. Responsiveness was not increased in response to iv MCh in vivo or EFS or MCh challenge in vitro. Increased epithelial permeability was not detected at 7 d.ConclusionInfection with 1 × 105 pfu RSV induced extreme hyperresponsiveness to aerosolised MCh during the acute phase of infection in adult and weanling mice. The route-specificity of hyperresponsiveness suggests that epithelial mechanisms were important in determining the physiological effects. Inflammatory changes were dissociated from physiological changes, particularly in weanling mice.

Age-Induced Reprogramming of Mast Cell Degranulation

Mast cell degranulation can initiate an acute inflammatory response and contribute to the progression of chronic diseases. Alteration in the cellular programs that determine the requirement for mast cell degranulation would therefore have the potential to dramatically impact disease severity. Mast cells are exposed to increased levels of PGE2 during inflammation. We show that although PGE2 does not trigger the degranulation of dermal mast cells of young animals, in older mice, PGE2 is a potent mast cell stimulator. Intradermal administration of PGE2 leads to an EP3 receptor-dependent degranulation of mast cells, with the number of degranulated cells approaching levels observed in IgE- and Ag-treated controls. Taken together, these studies suggest that the ability of PGE2 to initiate mast cell degranulation changes in the aging animal. Therefore, elevated PGE2 levels might provide an important pathway by which mast cells are engaged to participate in inflammatory responses in the elderly patient.

Immune parameters in patients with depression versus healthy controls during an open-label, placebo-controlled, randomized trial of reboxetine and celecoxib

Introduction: There is increasing evidence for the involvement of an immune process in the pathophysiology of major depression (MD). Previous studies could demonstrate elevated levels of proinflammatory cytokines as well as elevated PGE2 levels. Celecoxib is a COX–2 inhibitor, leading to a reduced production of PGE2. We conducted a randomized and placebo-controlled trial of celecoxib as add-on therapy in patients with MD treated with reboxetine.

Prostaglandin E2 Is a Product of Induced Prostaglandin-endoperoxide Synthase 2 and Microsomal-type Prostaglandin E Synthase at the Implantation Site of the Hamster

Certain uterine prostaglandins (PGs) are elevated at implantation sites and are needed to trigger the events of blastocyst implantation that include blastocyst-uterine attachment and stromal decidualization with vascular permeability changes. Several decades of investigations showed that treatment with PG synthesis inhibitors, prior to or during the time of implantation, resulted in either complete inhibition or a delay in implantation or reduction in the number of implantation sites with diminished decidual tissue. Consistent with these findings, we observed that whereas a selective PG endoperoxide synthase (Ptgs) 1 inhibitor SC-560 failed to inhibit implantation, a selective Ptgs2 inhibitor SC-236 showed significantly reduced number and size of implantation sites in progesterone-treated ovariectomized pregnant hamsters. It is known that Ptgs2 expression and Ptgs2-derived prostacyclin (PGI2) synthesis at implantation sites are needed for implantation in the mouse (a rodent that needs ovarian estrogen for implantation). However, it is unknown which Ptgs and PG synthases produce which PGs at implantation sites of the hamster (a rodent that does not need ovarian estrogen for implantation). Here we demonstrate that as blastocyst implantation proceeds, a reduction in Ptgs1 expression from uterine luminal epithelial cells and a gradual induction in Ptgs2 expression exclusively in luminal epithelial and adjacent decidual cells occurred at implantation sites of hamsters. Results also reveal that PGE2, but not PGI2, is the major PG at implantation sites where Ptgs2 and microsomal type PGE synthases but not PGI synthases are co-expressed. This elevated uterine PGE2 at implantation sites may serve to initiate or amplify physiological signals required for specific aspects of the implantation process in hamsters.

Targeted over-expression of mPGES-1 and elevated PGE2 production is not sufficient for lung tumorigenesis in mice.

There is a significant body of evidence suggesting that enzymes involved in arachidonic acid metabolism and their eicosanoid products play a role in various cancers, having both pro- and antitumorigenic effects. The goal of this study was to further define the role microsomal prostaglandin E synthases (mPGES-1) play in lung tumorigenesis. Transgenic mice were created with targeted over-expression of human mPGES-1 in the alveolar and airway epithelial cells using an SP-C promoter driven construct. Transgene positive (mPGES-1+) mice were shown to significantly over-express functional mPGES-1 in the lung and more specifically in alveolar type II cells. To study the effects of mPGES-1 over-expression in lung tumor formation, mice were exposed to a complete carcinogen protocol with a single injection of urethane or an initiation/promotion model with a single injection of 3-methylcholanthrene (MCA) followed by multiple injections of butylated hydroxytoluene (BHT). mPGES-1+ mice did not show a significant difference in tumor multiplicity or tumor size at 10, 16, 19 or 30 weeks after urethane injection compared with mPGES-1- mice. No significant difference was seen in tumor incidence, multiplicity or size at 19 weeks after treatment with MCA/BHT. Western blots verified that mPGES-1 expression was increased in tumors versus uninvolved tissue of both mPGES-1+ and mPGES-1- mice with overall expression being significantly higher in mPGES-1+ mice. Cyclooxygenase-2 levels were elevated in tumors in both groups. From these studies we conclude that over-expression of mPGES-1 and highly elevated PGE2 production are not sufficient to induce lung tumors.

Methanandamide increases COX-2 expression and tumor growth in murine lung cancer.

Increased COX-2 expression and elevated PGE2 have been associated with a poor prognosis in lung cancer. Cannabinoids have been known to exert some of their biological effects via modulation of prostaglandin production. We evaluated the impact of methanandamide on COX-2 expression, PGE2 production, and tumor growth in murine lung cancer. Methanandamide administration (5 mg/kg, four times/wk i.p.) resulted in an increased rate of tumor growth (P<0.01 compared with diluent treated controls). The CB1 and CB2 receptor antagonists, SR141716 and SR144528, did not block the methanandamide-mediated increase in tumor growth. In vivo, methanandamide treatment increased the production of PGE2 at the tumor site as well as in splenocytes. Consistent with these results, methanandamide increased PGE2 and COX-2 levels in murine lung cancer cells in vitro via a cannabinoid receptor-independent mechanism. The COX-2-specific inhibitor, SC58236, abrogated methanandamide induction of PGE2 production in vitro and blocked methanandamide-enhanced tumor growth in vivo. Furthermore, the p38/MAPK inhibitor, SB203528, and the p42/44 inhibitor, PD98059, blocked methanandamide-mediated induction of PGE2 and COX-2. These results suggest that methanandamide augments tumor growth by a cannabinoid receptor-independent pathway that is associated with the up-regulation of COX-2.