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Background. Overcoming radioresistance is an important problem in radiation oncology. Therefore, the development of new approaches to modeling the radiosensitivity of tumors in cancer patients becomes relevant and important. Cyclooxygenase-2 (COX-2) inhibitors are new agents for radiomodification in various radiation therapy schemes, the use of which slows down angiogenesis by suppressing the activity of the COX-2 enzyme. Purpose. To determine the content of indicators of radioresistance: vascular endothelial growth factor (VEGF), COX-2, prostaglandin E-2 (PGE-2) in the blood serum of patients with non-small cell lung cancer (NSCLC) and changes in their levels with different schemes of radiation therapy (RT). Materials and methods. 36 patients with NSCLC were examined and treated, who were divided into four groups: RT (the first group – 16 patients), RT with the COX-2 inhibitor – ranselex (the second group – 9 patients), RT with ranselex and cisplatin (the third group – 5 patients ) and RT with cisplatin (fourth group – 6 patients). The patients received a course of radiation treatment using a Clinac 600C linear accelerator. The classical fractionation mode was used, the total focal doses were 60–66 Gy. Cisplatin was prescribed at 30 mg/m2 per week up to a total dose (SD) of 200 mg, the COX-2 inhibitor Rancelex® at a dose of 100 mg per day (active substance – celecoxib). The levels of VEGF, COX-2, and PGE-2 in the blood serum of patients with NSCLC were determined by enzyme-linked immunosorbent assay (ELISA) before and after treatment. Results. The level of the angiogenesis marker VEGF after treatment in the group with RT decreases by 1.46 times, in the group with the combined action of RT and ranselex – 2.4 times, in the group with the combined action of PT, ranselex and cisplatin – by 3.7 times, and in the group with the combined effect of RT and cisplatin, it decreases by 1.1 times. The greatest decrease in the level of VEGF is observed with RT in combination with ranselex and cisplatin, which indicates a more effective enhancement of the antiangiogenic effect. Conclusions. It has been proven that with various schemes of RT using the COX-2 inhibitor – ranselex and cisplatin in patients with NSCLC, there is a decrease in the radioresistance markers PGE-2 and COX-2, the angiogenesis marker – VEGF, which indicates that the effect of radiomodification on the angiogenesis process is most pronounced in the combined actions of RT and both radio modifiers. The use of COX-2 inhibitors as radiosensitizers in combination with RT provides a new opportunity to increase tumor radiosensitivity., Актуальність. Важливою проблемою у радіаційній онкології є подолання радіорезистентності. Тому розробка нових підходів моделювання радіочутливості пухлин в онкологічних хворих стає актуальною та важливою. Новими агентами для радіомодифікації при різних схемах променевої терапії є інгібітори циклооксигенази-2 (ЦОГ-2), застосування яких призводить до уповільнення ангіогенезу шляхом пригнічення активності ферменту ЦОГ-2. Мета роботи. Визначити вміст показників радіорезистентності: фактор росту ендотелію судин (VEGF), ЦОГ-2 та простагландин Е-2 (ПГЕ-2) у сироватці крові хворих на недрібноклітинний рак легені (НДРЛ) та зміни їх рівнів при різних схемах променевої терапії (ПТ). Матеріали і методи. Обстежено та проліковано 36 хворих на НДРЛ, яких поділено на чотири групи: перша група – ПТ (16 хворих), друга група –ПТ з інгібітором ЦОГ-2 – ранселексом (9 хворих), третя група – ПТ з ранселексом та цисплатином (5 хворих) та четверта група – ПТ з цисплатином (6 хворих). Хворі отримували курс променевого лікування на лінійному прискорювачі «Clinac 600C». Використовувався режим класичного фракціонування, сумарні осередкові дози склали 60–66 Гр. Цисплатин призначали 30 мг/м2 на тиждень до сумарної дози 200 мг, інгібітор ЦОГ-2 – Ранселекс® – у дозі 100 мг на добу (діюча речовина – целекоксиб). Рівні VEGF, ЦОГ-2 та ПГЕ-2 у сироватці крові хворих на НДРЛ визначали методом імуноферментного аналізу (ІФА) до та після лікування. Результати. Рівень маркера ангіогенезу VEGF після лікування у першій групі при ПТ знижується у 1,46 рази, у групі при поєднаній дії ПТ та ранселексу – 2,4 рази, у групі при поєднаній дії ПТ, ранселексу та цисплатину – у 3,7 рази, а у групі при поєднаній дії ПТ та цисплатину знижується у 1,1 рази. Найбільше зниження рівня VEGF спостерігається при ПТ у поєднанні з ранселексом та цисплатином, що вказує на більш ефективне підсилення антиангіогенного ефекту. Висновки. Доведено, що при різних схемах ПТ з використанням інгібітора ЦОГ-2 – ранселексу, та цисплатину у хворих на НДРЛ спостерігається зниження маркерів радіорезистентності ПГЕ-2 та ЦОГ-2, маркера ангіогенезу – VEGF, це свідчить що вплив радіомодифікації на процес ангіогенезу найбільш виражений при поєднаній дії ПТ та обох радіомодифікаторів. Використання інгібіторів ЦОГ-2 як радіосенсибілізаторів у поєднанні з ПТ дає нову можливість підвищувати радіочутливість пухлини.

Nonsteroidal anti-inflammatory drugs are among the most commonly administered drugs in the perioperative period due to their prominent role in pain management. However, they potentially have perioperative consequences due to immune-modulating effects through the inhibition of prostanoid synthesis, thereby affecting the levels of various cytokines. These effects may have a direct impact on the postoperative outcome of patients since the immune system aims to restore homeostasis and plays an indispensable role in regeneration and repair. By affecting the immune response, consequences can be expected on various organ systems. This narrative review aims to highlight these potential immune system–related consequences, which include systemic inflammatory response syndrome, acute respiratory distress syndrome, immediate and persistent postoperative pain, effects on oncological and neurologic outcome, and wound, anastomotic, and bone healing.

Inflammatory response against implanted biomaterials impairs their functional integration and induces medical complications in the host's body. To suppress such immune responses, one approach is the administration of multiple drugs to halt inflammatory pathways. This challenges patient's adherence and can cause additional complications such as infection. Alternatively, biologics that regulate multiple inflammatory pathways are attractive agents in addressing the implants immune complications. Secretome of mesenchymal stromal cells (MSCs) is a multipotent biologic, regulating the homeostasis of lymphocytes and leukocytes. Here, it is reported that alginate microcapsules loaded with processed conditioned media (pCM-Alg) reduces the infiltration and/or expression of CD68+ macrophages likely through the controlled release of pCM. In vitro cultures revealed that alginate can dose dependently induce macrophages to secrete TNFα, IL-6, IL-1β, and GM-CSF. Addition of pCM to the cultures attenuates the secretion of TNFα (p = 0.023) and IL-6 (p

The immunomodulatory potential of mesenchymal stromal cells (MSCs) and regulatory T cells (T‐reg) is well recognized by translational scientists in the field of regenerative medicine and cellular therapies. A wide range of preclinical studies as well as a limited number of human clinical trials of MSC therapies have not only shown promising safety and efficacy profiles but have also revealed changes in T‐reg frequency and function. However, the mechanisms underlying this potentially important observation are not well understood and, consequently, the optimal strategies for harnessing MSC/T‐reg cross‐talk remain elusive. Cell‐to‐cell contact, production of soluble factors, reprogramming of antigen presenting cells to tolerogenic phenotypes, and induction of extracellular vesicles (“exosomes”) have emerged as possible mechanisms by which MSCs produce an immune‐modulatory milieu for T‐reg expansion. Additionally, these two cell types have the potential to complement each other's immunoregulatory functions, and a combinatorial approach may exert synergistic effects for the treatment of immunological diseases. In this review, we critically assess recent translational research related to the outcomes and mechanistic basis of MSC effects on T‐reg and provide a perspective on the potential for this knowledge base to be further exploited for the treatment of autoimmune disorders and transplants., Mesenchymal stromal cells (MSCs) promote regulatory T cell numbers and immune suppressive functions through mechanisms involving cell‐cell contact, production of soluble mediators, reprogramming of antigen presenting cells, and release of extracellular vesicles that likely represent important components of MSC therapeutic effects in immune/inflammatory diseases.

Most airway diseases, including chronic obstructive pulmonary disease (COPD), are associated with excessive coughing. The extent to which this may be a consequence of increased activation of vagal afferents by pathology in the airways (e.g., inflammatory mediators, excessive mucus) or an altered neuronal phenotype is unknown. Understanding whether respiratory diseases are associated with dysfunction of airway sensory nerves has the potential to identify novel therapeutic targets.To assess the changes in cough responses to a range of inhaled irritants in COPD and model these in animals to investigate the underlying mechanisms.Cough responses to inhaled stimuli in patients with COPD, healthy smokers, refractory chronic cough, asthma, and healthy volunteers were assessed and compared with vagus/airway nerve and cough responses in a cigarette smoke (CS) exposure guinea pig model.Patients with COPD had heightened cough responses to capsaicin but reduced responses to prostaglandin E2 compared with healthy volunteers. Furthermore, the different patient groups all exhibited different patterns of modulation of cough responses. Consistent with these findings, capsaicin caused a greater number of coughs in CS-exposed guinea pigs than in control animals; similar increased responses were observed in ex vivo vagus nerve and neuron cell bodies in the vagal ganglia. However, responses to prostaglandin E2 were decreased by CS exposure.CS exposure is capable of inducing responses consistent with phenotypic switching in airway sensory nerves comparable with the cough responses observed in patients with COPD. Moreover, the differing profiles of cough responses support the concept of disease-specific neurophenotypes in airway disease. Clinical trial registered with www.clinicaltrials.gov (NCT 01297790).

Background: Inflammatory and degenerative activity inside the joint can be studied in vivo via analysis of synovial fluid (SF) biomarkers, which are molecular markers of inflammatory processes and tissue turnover. The aim of this study was to investigate the response of selected biomarkers in the SF after an intra-articular (IA) high-molecular-weight non-animal stabilized hyaluronic acid (NASHA) treatment. Our hypothesis was that prostaglandin E-2 (PGE(2)), substance P, aggrecan chondroitin sulfate 846 epitope (CS846), and carboxypeptide of type II collagen (CPII) concentrations in SF would decrease more in the NASHA than in the placebo group. Twenty-eight clinically lame horses with positive responses to diagnostic IA anaesthesia of the metacarpophalangeal or metatarsophalangeal joints were randomized into treatment (n = 15) and control (n = 13) groups. After collection of baseline SF samples followed by IA diagnostic anaesthesia, horses in the treatment group received 3 ml of a NASHA product IA. Those in the placebo group received an equivalent volume of sterile 0.9% saline solution. The horses were re-evaluated and a second SF sample was obtained after a 2-week period. Results: CS846 concentration decreased in the NASHA group only (P = 0.010). Both PGE(2) and CPII concentrations decreased within the groups (PGE(2), P = 0.010 for the NASHA group; P = 0.027 for the placebo group; CPII, P

Objective: The Hoffa's fat pad (HFP) is an intra-articular adipose tissue which is situated under and behind the patella. It contains immune cells next to adipocytes and secretes inflammatory factors during osteoarthritis (OA). In this study, we compared the release profile of prostanoids, which are involved in inflammation, of HFP from OA patients vs patients with a focal cartilage defect (CD) without evidence for OA on MRI and investigated the prostanoid modulatory anti-inflammatory action of celecoxib on HFP. Design: Prostanoid release was analyzed in conditioned medium of HFP explant cultures from 17 osteoarthritic patients and 12 CD patients, in the presence or absence of celecoxib. Furthermore, gene expression of COX enzymes and expression of genes indicative of a pro-inflammatory or anti-inflammatory phenotype of HFP was analyzed. Results: Prostanoid release by HFP from knee OA patients clustered in two subgroups with high and low prostanoid producers. HFP from high prostanoid producers released higher amounts of PGE(2), PGF(2 alpha) and PGD(2) compared to HFP from CD patients. PGE(2) release by OA HFP was positively associated with expression of genes known to be expressed by M1 macrophages, indicating a role for macrophages. Celecoxib modulated prostanoid release by HFP, and also modulated the inflammation ratio towards a more favorable anti-inflammatory M2 phenotype, most effectively in patients with higher prostanoid release profiles. Conclusion: In knee OA patients with inflamed HFP's, celecoxib may exert positive effects in the knee joint via decreasing the release of prostanoids produced by the HFP and by favorably modulating the anti-inflammatory marker expression in HFP. (C) 2018 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

Summary Objective To define if adipose mesenchymal stromal cell (ASC) treatment mediated switching of the pro-inflammatory profile of M1-like macrophages as a means to develop a tailored in vitro efficacy/potency test. Design We firstly performed immunohistochemical analysis of CD68, CD80 (M1-like) and CD206 (M2-like) macrophages in osteoarthritic (OA) synovial tissue. ASC were co-cultured in contact and in transwell with activated (GM-CSF + IFNγ)-M1 macrophages. We analyzed IL1β, TNFα, IL6, MIP1α/CCL3, S100A8, S100A9, IL10, CD163 and CD206 by qRT-PCR or immunoassays. Prostaglandin E2 (PGE2) blocking experiments were performed using PGE2 receptor antagonist. Results In moderate grade OA synovium we did not always find a higher percentage of CD80 with respect to CD206. M1-like-activated macrophage factors IL1β, TNFα, IL6, MIP1α/CCL3, S100A8 and S100A9 were down-modulated both in contact and in transwell by ASC. However, in both systems ASC induced the typical M2-like macrophage markers IL10, CD163 and CD206. Activated-M1-like macrophages pre-treated with PGE2 receptor antagonist failed to decrease secretion of TNFα, IL6 and to increase that of IL10, CD163 and CD206 when co-cultured with ASC confirming a PGE2 specific role. Conclusions We demonstrated that ASC are responsible for the switching of activated-M1-like inflammatory macrophages to a M2-like phenotype, mainly through PGE2. This evidenced that activated-M1-like macrophages may represent a relevant cell model to test the efficacy/potency of ASC and suggests a specific role of ASC as important determinants in therapeutic dampening of synovial inflammation in OA.

International audience; Intra-articular drug delivery systems (DDSs) are envisaged as interesting alternative to locally release non-steroidal anti-inflammatory drugs (NSAIDs) such as ibuprofen to reduce pain in patients with osteoarthritis. The present study examines the efficacy of S-(+)-ibuprofen on cartilage degradation as drug candidate for DDS loading. Humeral cartilage and joint capsule explants were collected from healthy sheep shoulder joints and they were cultured in mono-or in co-culture for 13 days with LPS in combination with S-(+)-ibuprofen at 50 mu M and 1 mM. S-(+)-ibuprofen ( 50 mu M) blocked prostaglandins production in LPS-activated explants but did not reduce cartilage degradation. By contrast, 1mM S-(+)-ibuprofen treatment of cartilage explants reduced nitric oxide synthesis by 51% (p = 0.0072), proteoglycans degradation by 35% (p = 0.0114) and expression of serum amyloid protein -the main protein induced upon LPS challenge - by 44% (p

We have modeled the transition from acute to chronic pain in the rat. In this model (termed hyperalgesic priming) a chronic state develops after a prior inflammatory process or exposure to an inflammatory mediator, in which response to subsequent exposure to prostaglandin E2 (PGE2) is characterized by a protein kinase Cε-dependent marked prolongation of mechanical hyperalgesia. To assess the effect of priming on the function of the nociceptor, we have performed in vitro patch clamp and in vivo single-fiber electrophysiology studies using tumor necrosis factor α to induce priming. In vitro, the only change observed in nociceptors cultured from primed animals was a marked hyperpolarization in resting membrane potential (RMP); prolonged sensitization, measured at 60 minutes, could not be tested in vitro. However, complimentary with behavioral findings, in vivo baseline mechanical nociceptive threshold was significantly elevated compared to controls. Thirty minutes after injection of PGE2 into the peripheral receptive field, both primed and control nociceptors showed enhanced response to mechanical stimulation. However, 60 minutes after PGE2 administration, the response to mechanical stimulation was further increased in primed but not in control nociceptors. Thus, at the level of the primary afferent nociceptor, it is possible to demonstrate both altered function at baseline and prolonged PGE2-induced sensitization. Intrathecal antisense (AS) to Kv7.2, which contributes to RMP in sensory neurons, reversibly prevented the expression of priming in both behavioral and single-fiber electrophysiology experiments, implicating these channels in the expression of hyperalgesic priming.

Background: Recently, intra-articular botulinum toxin A (IA BoNT A) has been shown to reduce joint pain in osteoarthritic dogs. Similar results have been reported in human patients with arthritis. However, the mechanism of the antinociceptive action of IA BoNT A is currently not known. The aim of this study was to explore this mechanism of action by investigating the effect of IA BoNT A on synovial fluid (SF) and serum substance P (SP), prostaglandin E-2 (PGE(2)), and tumor necrosis factor alpha (TNF-alpha) in osteoarthritic dogs. Additionally, the aim was to compare SF SP and PGE(2) between osteoarthritic and non-osteoarthritic joints, and investigate associations between SP, PGE(2), osteoarthritic pain, and the signalment of dogs. Thirty-five dogs with chronic naturally occurring osteoarthritis and 13 non-osteoarthritic control dogs were included in the study. Osteoarthritic dogs received either IA BoNT A (n = 19) or IA placebo (n = 16). Serum and SF samples were collected and osteoarthritic pain was evaluated before (baseline) and 2 and 8 weeks after treatment. Osteoarthritic pain was assessed with force platform, Helsinki Chronic Pain Index, and joint palpation. Synovial fluid samples were obtained from control dogs after euthanasia. The change from baseline in SP and PGE(2) concentration was compared between the IA BoNT A and placebo groups. The synovial fluid SP and PGE(2) concentration was compared between osteoarthritic and control joints. Associations between SP, PGE(2), osteoarthritic pain, and the signalment of dogs were evaluated. Results: There was no significant change from baseline in SP or PGE(2) after IA BoNT A. Synovial fluid PGE(2) was significantly higher in osteoarthritic compared to control joints. Synovial fluid PGE(2) correlated with osteoarthritic pain. No associations were found between SP or PGE2 and the signalment of dogs. The concentration of TNF-alpha remained under the detection limit of the assay in all samples. Conclusions: The results suggest that the antinociceptive effect of IA BoNT A in the joint might not be related to the inhibition of SP nor PGE(2). Synovial fluid PGE(2,) but not SP, could be a marker for chronic osteoarthritis and pain in dogs.

Signal transduction by the cAMP/cAMP-dependent protein kinase A (PKA) pathway is triggered through multiple receptors and is important for many processes in a variety of cells. In T cells, the engagement of the TCR-CD3 complex induces CAMP, a second messenger that controls immune response. IL-10, produced by a variety of lymphocyte subpopulations, is an important regulator of this response exerting a wide range of immunomodulatory actions. Elevation of cAMP has been shown to increase IL-10 production by monocytes. However, the mechanism of cAMP mediated regulation of IL-10 production by T lymphocytes remains unclear. In this study using normal peripheral T lymphocytes stimulated either through the TCR-CD3 complex or the TCR-CD3 and the CD28 molecule, we show that IL-10 is produced mainly by memory T lymphocytes after either way of stimulation and is drastically inhibited (70-90%) by CAMP elevating agents. cAMP mediated inhibition was reversed by the use of the specific PKA inhibitor Rp-8-Br-cAMP but not by the addition of exogenous rhIL-2, indicating that the inhibitory effect depends on PKA activation and is not secondary to IL-2 inhibition, Inhibition is taking place at both transcriptional and posttranscriptional level. Transfection of a luciferase reporter plasmid carrying the IL-10 promoter in T cells, revealed that TCR/CD28-induced activation was inhibited by 60% by cAMP elevation. The most sensitive part to cAMP mediated inhibition was a fragment of 135 bp Upstream of TATA box, which contains multiple binding sites for MEF-2. Overexpression of MEF-2 in the same cells increased IL-10 promoter activity by 2.5-fold. Stimulation through TCR/CD28 increased MEF-2 binding in its Corresponding binding sites which was inhibited by 80% in the presence of cAMP elevating agents. These results suggest that the inhibitory effect of cAMP on IL-10 production by normal peripheral T lymphocytes is cell type and stimulus specific, exerted on multiple levels and involves MEF2 transcription factor. (C) 2008 Elsevier Ltd. All rights reserved. Mol Immunol

The innate immune response to bacterial infections requires the interaction of neutrophils and platelets. Here, we show that a multistep reciprocal crosstalk exists between these two cell types, ultimately facilitating neutrophil influx into the lung to eliminate infections. Activated platelets adhere to intravascular neutrophils through P-selectin/P-selectin glycoprotein ligand-1 (PSGL-1)-mediated binding, a primary interaction that allows platelets glycoprotein Ibα (GPIbα)-induced generation of neutrophil-derived extracellular vesicles (EV). EV production is directed by exocytosis and allows shuttling of arachidonic acid into platelets. EVs are then specifically internalized into platelets in a Mac1-dependent fashion, and relocated into intracellular compartments enriched in cyclooxygenase1 (Cox1), an enzyme processing arachidonic acid to synthesize thromboxane A2 (TxA2). Finally, platelet-derived-TxA2 elicits a full neutrophil response by inducing the endothelial expression of ICAM-1, intravascular crawling, and extravasation. We conclude that critical substrate–enzyme pairs are compartmentalized in neutrophils and platelets during steady state limiting non-specific inflammation, but bacterial infection triggers regulated EV shuttling resulting in robust inflammation and pathogen clearance., Interaction between platelets and neutrophils promotes neutrophil activation. Here the authors show that neutrophils initiate the cross-talk with platelets by shuttling arachidonic acid via extracellular vesicles, which platelets convert to thromboxane A2 that then elicits neutrophil activation.

Fos expression was used to assess whether the proinflammatory cytokine interleukin-1beta (IL-1beta) activated specific, chemically coded neuronal populations in isolated preparations of guinea pig ileum and colon. Whether the effects of IL-1beta were mediated through a prostaglandin pathway and whether IL-1beta induced the expression of cyclooxygenase (COX)-2 was also examined. Single- and double-labeling immunohistochemistry was used after treatment of isolated tissues with IL-1beta (0.1-10 ng/ml). IL-1beta induced Fos expression in enteric neurons and also in enteric glia in the ileum and colon. For enteric neurons, activation was concentration-dependent and sensitive to indomethacin, in both the myenteric and submucosal plexuses in both regions of the gut. The maximum proportion of activated neurons differed between the ileal (approximately 15%) and colonic (approximately 42%) myenteric and ileal (approximately 60%) and colonic (approximately 75%) submucosal plexuses. The majority of neurons activated in the myenteric plexus of the ileum expressed nitric oxide synthase (NOS) or enkephalin immunoreactivity. In the colon, activated myenteric neurons expressed NOS. In the submucosal plexus of both regions of the gut, the majority of activated neurons were vasoactive intestinal polypeptide (VIP) immunoreactive. After treatment with IL-1beta, COX-2 immunoreactivity was detected in the wall of the gut in both neurons and nonneuronal cells. In conclusion, we have found that the proinflammatory cytokine IL-1beta specifically activates certain neurochemically defined neural pathways and that these changes may lead to disturbances in motility observed in the inflamed bowel.

Background: Beta-sitosterol (β-sitosterol) is one of the several phytosterols widely studied for its potential to reduce benign prostatic hyperplasia and blood cholesterol levels. Objective: In the present study, the effects of β-sitosterol on spontaneous and agonist-induced contractions in in vitro nonpregnant human uterus with respect to prostaglandin E2(PGE2) were investigated. Materials and Methods: Myometrial strips, measuring approximately 15 mm × 4 mm × 2 mm, were attained from hysterectomy samples of premenopausal women. Longitudinal muscle strips were mounted on tissue baths, under physiological conditions, to measure their isometric contraction. The effects of cumulative amounts of β-sitosterol on spontaneous motility in the absence and presence of prazosin, atropine, fulvestran, indomethacin, or ethylenediaminetetraacetic acid (EDTA), and on agonist-induced motor activity, were examined. Results: On strips in the follicular phase, both β-sitosterol (1–100 μg/ml) and PGE2(0.1–10 μg/ml) increase, in a concentration-dependent manner, muscular basic tonus and amplitude and frequency of spontaneous uterine contractions; whereas on strips obtained during periovulatory phase, β-sitosterol and PGE2cause inhibition of uterine motility. For contractile response, the effective concentrations (EC50) were 47.8 μg/ml and 5.19 μg/ml, respectively. Unlike indomethacin, the tissue pretreatment with prazosin, fulvestran, atropine, or ethylenediaminetetraacetic acid did not affect the contractile uterine responses to β-sitosterol. Furthermore, the β-sitosterol was able to potentiate the contractile response induced by acetylcholine and vasopressin. Conclusions: These observations suggest that β-sitosterol may be a useful modulator of the uterine motility during menstrual cycle, facilitating female fertility.

Chronic obstructive pulmonary disease (COPD) is a progressive lung disease and is currently the fourth leading cause of death worldwide. Chronic inflammation and repair processes in the small airways are characteristic of COPD. Despite extensive efforts from researchers and industry, there is still no cure for COPD, hence an urgent need for new therapeutic alternatives. MicroRNAs are such an option; they are small noncoding RNAs involved in gene regulation. Their importance has been shown with respect to maintaining the balance between health and disease. Although previous reviews have discussed the expression of microRNAs related to lung disease, a detailed discussion regarding the function of differential miRNA expression in the pathogenesis of COPD is lacking.In this review we link the expression of microRNAs to different features of COPD and explain their importance in the pathogenesis of this disease. We further discuss their potential to contribute to the development of future therapeutic strategies.

The present study was undertaken to evaluate the effects of the glucocorticoid hormones betamethasone and hydrocortisone, and of progesterone on the relative production of prostaglandin E2 (PGE2) and leukotriene B4 (LTB4) by explants of human fetal membranes at term gestation in the absence of labor. Tissues (n = 7) were incubated either in the presence or in the absence of the above mentioned hormones. PGE2 and LTB4 were measured in culture medium by radioimmunoassays. Glucocorticoids and progesterone did not affect PGE2 output by tissues; however, they greatly stimulated LTB4 production. Moreover, both betamethasone and hydrocortisone significantly increased the ratio of LTB4 to PGE2 formation by tissues. These results suggest that glucocorticoid hormones and progesterone might influence arachidonic acid metabolism in human fetal membranes by stimulating the production of lipoxygenase rather than cyclooxygenase substances before the onset of labor.

Prostaglandins are synthesized from arachidonic acid by the enzyme cyclo-oxygenase. There are two isoforms of cyclooxygenases: COX-1 (a constitutive form) and COX-2 (an inducible form). COX-2 has recently been categorized as an immediate-early gene and is associated with cellular growth and differentiation. The purpose of this study was to investigate the effects of exogenous dimethylprostaglandin E2 (dmPGE2) on prostate cancer cell growth. Results of these experiments demonstrate that administration of dmPGE2 to growing PC-3 cells significantly increased cellular proliferation (as measured by the cell number), total DNA content and endogenous PGE2 concentration. DmPGE2 also increased the steady-state mRNA levels of its own inducible synthesizing enzyme, COX-2, as well as cellular growth to levels similar to those seen with fetal calf serum and phorbol ester. The same results were observed in other human cancer cell types, such as the androgen-dependent LNCaP cells, breast cancer MDA-MB-134 cells and human colorectal carcinoma DiFi cells. In PC-3 cells, the dmPGE2 regulation of the COX-2 mRNA levels was both time dependent, with maximum stimulation seen 2 h after addition, and dose dependent on dmPGE2 concentration, with maximum stimulation seen at 5 microg ml(-1). The non-steroidal anti-inflammatory drug flurbiprofen (5 microM), in the presence of exogenous dmPGE2, inhibited the up-regulation of COX-2 mRNA and PC-3 cell growth. Taken together, these data suggest that PGE2 has a specific role in the maintenance of human cancer cell growth and that the activation of COX-2 expression depends primarily upon newly synthesized PGE2, perhaps resulting from changes in local cellular PGE2 concentrations. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7

Highlights • We report interactions of Toll-like receptors (TLRs) with COX enzymes in vivo. • COX-2 was broadly induced by LPS (TLR4) but more locally by poly(I:C) (TLR3). • COX-1/2 deletion modified the response to TLR activation in a TLR-specific manner. • COX-2 deletion enhanced interferon responses to viral-type TLR3/7/9 ligands. • COX-2 inhibition could provide a novel anti-viral therapeutic strategy., Cyclooxygenase 2 (COX)-2 is induced by bacterial and viral infections and has complex, poorly understood roles in anti-pathogen immunity. Here, we use a knock-in luciferase reporter model to image Cox2 expression across a range of tissues in mice following treatment with the either the prototypical bacterial pathogen-associated molecular pattern (PAMP), LPS, which activates Toll-like receptor (TLR)4, or with poly(I:C), a viral PAMP, which activates TLR3. LPS induced Cox2 expression in all tissues examined. In contrast, poly(I:C) elicited a milder response, limited to a subset of tissues. A panel of cytokines and interferons was measured in plasma of wild-type, Cox1−/− and Cox2−/− mice treated with LPS, poly(I:C), MALP2 (TLR2/6), Pam3CSK4 (TLR2/1), R-848 (TLR7/8) or CpG ODN (TLR9), to establish whether/how each COX isoform modulates specific PAMP/TLR responses. Only LPS induced notable loss of condition in mice (inactivity, hunching, piloerection). However, all TLR agonists produced cytokine responses, many of which were modulated in specific fashions by Cox1 or Cox2 gene deletion. Notably we observed opposing effects of Cox2 gene deletion on the responses to the bacterial PAMP, LPS, and the viral PAMP, poly(I:C), consistent with the differing abilities of the PAMPs to induce Cox2 expression. Cox2 gene deletion limited the plasma IL-1β and interferon-γ responses and hypothermia produced by LPS. In contrast, in response to poly(I:C), Cox2−/− mice exhibited enhanced plasma interferon (IFNα,β,γ,λ) and related cytokine responses (IP-10, IL-12). These observations suggest that a COX-2 selective inhibitor, given early in infection, may enhance and/or prolong endogenous interferon responses, and thereby increase anti-viral immunity.

Despite studies demonstrating that inhibition of cyclooxygenase-2 (COX-2)-derived prostaglandin E2 (PGE2) has significant chemotherapeutic benefits in vitro and in vivo, inhibition of COX enzymes is associated with serious gastrointestinal and cardiovascular side effects, limiting the clinical utility of these drugs. PGE2 signals through four different receptors (EP1-EP4) and targeting individual receptor(s) may avoid these side effects, while retaining significant anticancer benefits. Here, we show that targeted inhibition of the EP1 receptor in the tumor cells and the tumor microenvironment resulted in the significant inhibition of tumor growth in vivo. Both dietary administration and direct injection of the EP1 receptor-specific antagonist, ONO-8713, effectively reduced the growth of established CT26 tumors in BALB/c mice, with suppression of the EP1 receptor in the tumor cells alone less effective in reducing tumor growth. This antitumor effect was associated with reduced Fas ligand expression and attenuated tumor-induced immune suppression. In particular, tumor infiltration by CD4+CD25+Foxp3+ regulatory T cells was decreased, whereas the cytotoxic activity of isolated splenocytes against CT26 cells was increased. F4/80+ macrophage infiltration was also decreased; however, there was no change in macrophage phenotype. These findings suggest that the EP1 receptor represents a potential target for the treatment of colon cancer.

Background Cudrania tricuspidata extract is an important traditional herbal remedy for tumors, inflammation, gastritis, and liver damage and is predominantly used in Korea, China, and Japan. However, the anti-inflammatory effects of the extract have not yet been conclusively proved. Methods In this study, we investigated the effects of the CHCl3 fraction (CTC) of a methanol extract of C. tricuspidata on the lipopolysaccharide (LPS)-induced nitric oxide (NO) and prostaglandin E2 (PGE2) production in RAW 264.7 macrophage cells and mouse peritoneal macrophages, and the levels of pro-inflammatory cytokines tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-6 in RAW 264.7 macrophage cells. Results We observed that the protein expression levels of inducible NO synthase and COX-2 enzymes were markedly inhibited by CTC in a concentration-dependent manner. In addition, CTC reduced the production of TNF-α, IL-1β, and IL-6 in the LPS-stimulated RAW 264.7 macrophage cells. Conclusions Our results show that the C. tricuspidata extract could modulate macrophage-mediated inflammatory functions such as the overproduction of cytokines, NO, and PGE2. The CTC was found to be the active fraction in this context.

The potential for dietary supplementation with n-3 polyunsaturated fatty acids (n-3 PUFA) to improve reproductive efficiency in cattle has received much interest. The mechanisms by which n-3 PUFA may affect physiological and biochemical processes in key reproductive tissues are likely to be mediated by significant alterations in gene expression. The objective of this study was to examine the effects of dietary n-3 PUFA supplementation on global uterine endometrial gene expression in cattle. Beef heifers were supplemented with a rumen protected source of either a saturated fatty acid (CON; palmitic acid) or high n-3 PUFA (n-3 PUFA; 275 g) diet per animal per day for 45 days and global gene expression was determined in uterine endometrial tissue using an Affymetrix oligonucleotide bovine array. A total of 1,807 (946 up- and 861 downregulated) genes were differentially expressed following n-3 PUFA supplementation. Dietary n-3 PUFA altered numerous cellular processes potentially important in the control of reproduction in cattle. These included prostaglandin biosynthesis, steroidogenesis and transcriptional regulation, while effects on genes involved in maternal immune response and tissue remodeling were also observed. This study provides new insights into the effects of n-3 PUFA supplementation on the regulation of gene expression in the bovine uterus.

We have previously found in human renal proximal tubular HK-2 cells that hypoxia- and all-trans retinoic acid-induced hypoxia-inducible factor-1 alpha up-regulation is accompanied by retinoic acid receptor-beta up-regulation. Here we first investigated whether hypoxia-inducible factor-1 alpha expression is dependent on retinoic acid receptor-beta and our results confirmed it since (i) hypoxia-inducible factor-1 alpha-inducing agents hypoxia, hypoxia-mimetic agent desferrioxamine, all-trans retinoic acid and interleukin-1 beta increased retinoic acid receptor-beta expression, (ii) hypoxia-inducible factor-1 alpha up-regulation was prevented by retinoic acid receptor-beta antagonist LE-135 or siRNA retinoic acid receptor-beta and (iii) there was direct binding of retinoic acid receptor-beta to the retinoic acid response element in hypoxia-inducible factor-1 alpha promoter upon treatment with all-trans retinoic acid and 16,16-dimethyl-prostaglandin E-2. Since intracellular prostaglandin E-2 mediates hypoxia-inducible factor-1 alpha up-regulation in normoxia in HK-2 cells, we next investigated and confirmed, its role in the up-regulation of retinoic acid receptor-beta in normoxia by hypoxia-inducible factor-lot-inducing agents all-trans retinoic acid, interleukin-1 beta and 16,16-dimethyl-prostaglandin E-2 by inhibiting cyclooxygenases, prostaglandin influx transporter or EP receptors. Interestingly, the hypoxia-induced increase in retinoic acid receptor-beta expression and accumulation of hypoxia-inducible factor-1 alpha was also blocked by the inhibitors tested. This is the first time, to our knowledge, that retinoic acid receptor-beta signalling is involved in the control of the expression of transcription factor hypoxia-inducible factor-la in both normoxia and hypoxia and that retinoic acid receptor-beta expression is found to be strictly regulated by intracellular prostaglandin E-2. Given the relevance of hypoxia-inducible factor-la in the kidney in terms of tumorigenesis, progressive renal failure, production of erythropoietin and protection in several models of renal disease, our results open new therapeutic opportunities on the control of hypoxia-inducible factor-la based upon the pharmacological modulation of retinoic acid receptor-beta, either directly or through the control of intracellular prostaglandin E-2 levels/signalling. (C) 2012 Elsevier Ltd. All rights reserved.

Glucosamine (GlcN) is used in the treatment of osteoarthritis as symptomatic slow-acting drug, but its mode of action is not completely known. We analyzed the influence of GlcN and its N-acetyl-phenylalanine derivative (NAPA) on mRNA transcription level of TNF-alpha-stimulated genes in cell culture.Human immortalized chondrocyte cell line lbpva55 was stimulated with TNF-alpha and treated with GlcN and NAPA. mRNA transcription level of several genes, identified by complementary DNA microarray (cDNA microarray), was validated by Quantitative Real-Time Polymerase Chain Reaction (Q-RT-PCR).Several genes, whose mRNA level was increased by TNF-alpha treatment and significantly reduced by GlcN and NAPA in lbpva55 cells, were identified. These include cytokine receptors TNF-R1 and TNF-R2, their associated factor TRAF-6, signaling intermediates IGFB-6 and Rnd1, as well as cell cycle regulating proteins CUL-2 and G1S protein 1. Down- regulation of mRNA expression level of some of these genes is in accordance with inactivation of NF-kB transcription factor. Moreover, we found down-regulation of c-jun mRNA level, a component of AP-1 transcription factor.Our study suggests that GlcN and NAPA interfere with activation of NF-kB and AP-1 transcription factors, which are responsible for the expression of genes involved in diverse biological processes, such as cell growth and death, inflammatory and stress responses, accounting for the beneficial effects of GlcN in osteoarthritis.

P388D1 cells release arachidonic acid (AA) and produce prostaglandin E2 (PGE2) upon long-term stimulation with lipopolysaccharide (LPS). The cytosolic Group IVA (GIVA) phospholipase A2 (PLA2) has been implicated in this pathway. LPS stimulation also results in increased expression and secretion of a secretory PLA2, specifically GV PLA2. To test whether GV PLA2 contributes to PGE2 production and whether GIVA PLA2 activation increases the expression of GV PLA2, we utilized the specific GIVA PLA2 inhibitor pyrrophenone and second generation antisense oligonucleotides (AS-ONs) designed to specifically inhibit expression and activity of GV PLA2. Treatment of P388D1 cells with antisense caused a marked decrease in basal GV PLA2 mRNA and prevented the LPS-induced increase in GV PLA2 mRNA. LPS-stimulated cells release active GV PLA2 into the medium, which is inhibited to background levels by antisense treatment. However, LPS-induced PGE2 release by antisense-treated cells and by control cells are not significantly different. Collectively, the results suggest that the upregulation of GV PLA2 during long-term LPS stimulation is not required for PGE2 production by P388D1 cells. Experiments employing pyrrophenone suggested that GIVA PLA2 is the dominant player involved in AA release, but it appears not to be involved in the regulation of LPS-induced expression of GV PLA2 or cyclooxygenase-2.

Background: Protein kinase A (PKA) activation is documented to be inhibitory for T helper cell (T-H1)-like cytokines (IL-2, IFN-gamma), whereas T-H2-like cytokines (IL-4, IL-5) are not affected or upregulated. We have recently shown that IL-4 gene expression can be inhibited by PKA activation but depends on the mode of T-cell activation. For IL-5 gene expression, we hypothesized that the mode of T-cell activation also determines the ultimate effect of simultaneous PKA activation. Objectives: The objective of this study was the examination of IL-5 gene expression in healthy T cells activated with various mitogenic stimuli after simultaneous activation of PKA by dibutyryl-cAMP or prostaglandin E-2 (PGE(2)). Methods: IL-5 mRNA was measured by semiquantitative reverse transcription-polymerase chain reaction or Northern analysis. IL-5 protein was measured by ELISA. Transcriptional mechanisms involved in IL-5 gene expression were determined by nuclear run-on experiments and electrophoretic mobility shift assays. Posttranscriptional mechanisms were determined by actinomycin D chase studies. Results: Anti-CD2-, anti-CD3-, and anti-CD3 plus anti-CD28-induced IL-5 mRNA were completely inhibited by dibutyryl cyclic AMP (10(-3) mol/L) and PGE(2) (10(-5) mol/L), whereas concanavalin A-induced IL-5 mRNA was partially reduced. The effect of PGE(2) was accomplished at the transcriptional level, probably as the result of inhibition of DNA binding of nuclear factor-kappa B. Anti-CD3 plus anti-CD28-induced IL-5 protein (504 +/- 56 pg/ml) was significantly reduced by PGE(2) (122 +/- 42 pg/ml, p