Diosgenin, a plant steroid, induces apoptosis in human rheumatoid arthritis synoviocytes with cyclooxygenase-2 overexpression

In the present study, we have shown for the first time that a plant steroid, diosgenin, causes an inhibition of the growth of fibroblast-like synoviocytes from human rheumatoid arthritis, with apoptosis induction associated with cyclooxygenase-2 (COX-2) up-regulation. Celecoxib, a selective COX-2 inhibitor, provoked a large decrease in diosgenin-induced apoptosis even in the presence of exogenous prostaglandin E(2), whereas interleukin-1β, a COX-2 inducer, strongly increased diosgenin-induced apoptosis of these synoviocytes. These findings suggest that the proapoptotic effect of diosgenin is associated with overexpression of COX-2 correlated with overproduction of endogenous prostaglandin E(2). We also observed a loss of mitochondrial membrane potential, caspase-3 activation, and DNA fragmentation after diosgenin treatment

Staphylococcus aureus enterotoxin B regulates prostaglandin E-2 synthesis, growth, and migration in nasal tissue fibroblasts

Background. Superantigens and eicosanoids are important amplifiers and regulators of inflammation in airway diseases. We therefore studied the possible influence of Staphylococcus aureus enterotoxin B ( SEB) on the cyclooxygenase ( COX) pathway and basic functions of airway structural cells. Methods. Fibroblasts were isolated from nasal inferior turbinate tissue and cultured in the presence of different concentrations of SEB. Preincubation with interferon ( IFN)-gamma was performed to induce expression of major histocompatibility complex ( MHC) class II receptors. Prostaglandin E2 ( PGE(2)) production was assayed by enzyme-linked immunosorbent assay, and levels of COX-2 and prostanoid E receptors 1-4 ( EP1-4) were assayed by real-time polymerase chain reaction. Migration and growth tests were performed, and SEB was localized within the cells by confocal microscopy. Results. Stimulation with IFN-gamma and SEB significantly down-regulated PGE2, COX-2, and EP2 expression but not EP1, EP3, or EP4 expression. The enterotoxin blocked cell growth but increased the fibroblast migration rate. SEB was localized within the cell in the presence and absence of MHC-II, suggesting that mechanisms other than conventional binding may allow the enterotoxin to enter the cell. Conclusions. These findings may have major implications for our understanding of the role played by bacterial superantigens in regulating the inflammatory and remodeling mechanisms of upper airway diseases and hence may help elucidate the pathophysiology of these diseases

Activation of α7 nicotinic acetylcholine receptor by nicotine selectively up-regulates cyclooxygenase-2 and prostaglandin E(2 )in rat microglial cultures

BACKGROUND: Nicotinic acetylcholine (Ach) receptors are ligand-gated pentameric ion channels whose main function is to transmit signals for the neurotransmitter Ach in peripheral and central nervous system. However, the α7 nicotinic receptor has been recently found in several non-neuronal cells and described as an important regulator of cellular function. Nicotine and ACh have been recently reported to inhibit tumor necrosis factor-α (TNF-α) production in human macrophages as well as in mouse microglial cultures. In the present study, we investigated whether the stimulation of α7 nicotinic receptor by the specific agonist nicotine could affect the functional state of activated microglia by promoting and/or inhibiting the release of other important pro-inflammatory and lipid mediator such as prostaglandin E(2). METHODS: Expression of α7 nicotinic receptor in rat microglial cell was examined by RT-PCR, immunofluorescence staining and Western blot. The functional effects of α7 receptor activation were analyzed in resting or lipopolysaccharide (LPS) stimulated microglial cells pre-treated with nicotine. Culture media were assayed for the levels of tumor necrosis factor, interleukin-1β, nitric oxide, interleukin-10 and prostaglandin E(2). Total RNA was assayed by RT-PCR for the expression of COX-2 mRNA. RESULTS: Rat microglial cells express α7 nicotinic receptor, and its activation by nicotine dose-dependently reduces the LPS-induced release of TNF-α, but has little or no effect on nitric oxide, interleukin-10 and interleukin-1β. By contrast, nicotine enhances the expression of cyclooxygenase-2 and the synthesis of one of its major products, prostaglandin E(2). CONCLUSIONS: Since prostaglandin E(2 )modulates several macrophage and lymphocyte functions, which are instrumental for inflammatory resolution, our study further supports the existence of a brain cholinergic anti-inflammatory pathway mediated by α7 nicotinic receptor that could be potentially exploited for novel treatments of several neuropathologies in which local inflammation, sustained by activated microglia, plays a crucial role

The standardized herbal combination BNO 2103 contained in Canephron® N alleviates inflammatory pain in experimental cystitis and prostatitis

Background: Urinary tract infections are among the most common types of infections and give rise to inflammation with pain as one of the main symptoms. The herbal medicinal product Canephrod (R) N contains BNO 2103, a defined mixture of pulverized rosemary leaves, centaury herb, and lovage root, and has been used in the treatment of urinary tract infections for more than 25 years.Purpose: To test the hypothesis that BNO 2103 reduces pain in cystitis and prostatitis by virtue of anti-inflammatory properties, and to reveal potential mechanisms underlying the anti-inflammatory features.Study design: BNO 2103 was studied for anti-inflammatory and analgesic properties in three animal models in vivo, and the mode of action underlying the anti-inflammatory features was investigated in human leukocytes and cell-free assays in vitro.Methods: To assess the anti-inflammatory and analgesic efficacy of BNO 2103 we employed cyclophosphamide-induced cystitis and carrageenan-induced prostatitis in rats, and zymosan-induced peritonitis in mice. Human neutrophils and monocytes as well as isolated human 5-lipoxygenase and microsomal prostaglandin E-2 synthase-1-containing microsomes were utilized to assess inhibition of leukotriene and/or prostaglandin E-2 production by HPLC and/or ELISA.Results: When given orally, BNO 2103 reduced inflammation and hyperalgesia in experimental cystitis in rats, while individual components of BNO 2103 also reduced hyperalgesia. Furthermore, BNO 2103 reduced hyperalgesia in rats with carrageenan-induced prostatitis. Cell-based and cell-free studies implicate inhibition of prostaglandin E-2 and leukotriene B-4 biosynthesis as potential mechanisms underlying the analgesic and anti-inflammatory effects.Conclusion: Our data support the hypothesis that BNO 2103 reduces pain by virtue of its anti-inflammatory properties, possibly related to suppression of prostaglandin E-2 and leukotriene B-4 formation, and suggest that this combination has the potential to treat clinical symptoms such as inflammatory pain. Thus BNO 2103 may represent an alternative to reduce the use of antibiotics in urinary tract infections

Inducible nitric oxide synthase links NF-κB to PGE(2 )in polyunsaturated fatty acid altered fibroblast in-vitro wound healing

BACKGROUND: This study investigated mechanisms of altered fibroblast collagen production induced by polyunsaturated fatty acids. 3T3-Swiss fibroblasts were grown in medium containing either eicosapentaenoic or arachidonic acid. The effects of nuclear factor-kappaB activation by lipopolysaccharide on inducible nitric oxide synthase, nitric oxide, prostaglandin E(2), collagen production, and in-vitro wound healing were studied. RESULTS: Eicosapentaenoic acid treated cells produced less prostaglandin E(2 )but had increased inducible nitric oxide synthase expression, nitric oxide production, collagen formation, and recoverage area during in-vitro wound healing than cells treated with arachidonic acid. Activation of nuclear factor-kappaB with lipopolysaccharide increased inducible nitric oxide synthase expression, the production of nitric oxide, prostaglandin E(2), collagen, and the in-vitro wound recoverage area. The nitric oxide synthase inhibitor, N(G)-nitro-L-arginine methyl ester, decreased lipopolysaccharide-induced nitric oxide, but the amount of nitric oxide was greater in eicosapentaenoic acid treated cells. N(G)-nitro-L-arginine methyl ester plus lipopolysaccharide treatment increased collagen production and cellular recoverage area while treatment with N(G)-nitro-L-arginine methyl ester alone decreased it in wounded fibroblasts. CONCLUSION: The activation of the NF-κB pathway and PGE(2 )can be linked by the cross-talk of iNOS and NO in the PUFA altered fibroblast collagen production and wound healing. Additional studies are needed to determine how polyunsaturated fatty acids can be used as adjuvants in combination with other treatments (i.e, drugs) to design therapies to either enhance healthy collagen production or inhibit production and reduce fibrosis

Inhibition of release of inflammatory mediators in primary and cultured cells by a Chinese herbal medicine formula for allergic rhinitis

BACKGROUND: We demonstrated that a Chinese herbal formula, which we refer to as RCM-101, developed from a traditional Chinese medicine formula, reduced nasal and non-nasal symptoms of seasonal allergic rhinitis (SAR). The present study in primary and cultured cells was undertaken to investigate the effects of RCM-101 on the production/release of inflammatory mediators known to be involved in SAR. METHODS: Compound 48/80-induced histamine release was studied in rat peritoneal mast cells. Production of leukotriene B(4 )induced by the calcium ionophore A23187 was studied in porcine neutrophils using an HPLC assay and lipopolysaccharide-stimulated prostaglandin E(2 )production was studied in murine macrophage (Raw 264.7) cells by immune-enzyme assay. Expression of cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) was determined in Raw 264.7 cells, using western blotting techniques. RESULTS: RCM-101 (1–100 μg/mL) produced concentration-dependent inhibition of compound 48/80-induced histamine release from rat peritoneal mast cells and of lipopolysaccharide-stimulated prostaglandin E(2 )release from Raw 264.7 cells. Over the range 1 – 10 μg/mL, it inhibited A23187-induced leukotriene B(4 )production in porcine neutrophils. In addition, RCM-101 (100 μg/mL) inhibited the expression of COX-2 protein but did not affect that of COX-1. CONCLUSION: The findings indicate that RCM-101 inhibits the release and/or synthesis of histamine, leukotriene B(4 )and prostaglandin E(2 )in cultured cells. These interactions of RCM-101 with multiple inflammatory mediators are likely to be related to its ability to reduce symptoms of allergic rhinitis

N-Acetylcysteine enhances the action of anti-inflammatory drugs as suppressors of prostaglandin production in monocytes.

The anti-inflammatory effect of non-steroidal anti-inflammatory drugs (NSAIDs) is associated with inhibition of cyclooxygenase (COX), the rate-limiting enzyme responsible for the synthesis of prostaglandins. Since oxygen free radicals can act as second cellular messengers, especially to modulate the metabolism of arachidonic acid and the prostaglandin tract, it seems plausible that antioxidants might affect the production of prostaglandin by activated cells. This research is focused on the effect of the antioxidant N-acetylcysteine (NAC) on the inhibition of prostaglandin E(2) formation in activated monocytes by specific and non-specific COX inhibitors. We found that lipopolysaccharide-induced prostaglandin E(2) formation was significantly reduced by rofecoxib and by diclofenac, two NSAIDs. Addition of NAC to each of these drugs enhanced the effect of the NSAIDs. These results suggest that one might expect either a potentiation of the anti-inflammatory effect of COX inhibitors by their simultaneous administration with NAC, or obtaining the same anti-inflammatory at lower drug levels

Cyclooxygenases and prostaglandin E(2 )receptors in growth plate chondrocytes in vitro and in situ – prostaglandin E(2 )dependent proliferation of growth plate chondrocytes

Prostaglandin E(2 )(PGE(2)) plays an important role in bone development and metabolism. To interfere therapeutically in the PGE(2 )pathway, however, knowledge about the involved enzymes (cyclooxygenases) and receptors (PGE(2 )receptors) is essential. We therefore examined the production of PGE(2 )in cultured growth plate chondrocytes in vitro and the effects of exogenously added PGE(2 )on cell proliferation. Furthermore, we analysed the expression and spatial distribution of cyclooxygenase (COX)-1 and COX-2 and PGE(2 )receptor types EP1, EP2, EP3 and EP4 in the growth plate in situ and in vitro. PGE(2 )synthesis was determined by mass spectrometry, cell proliferation by DNA [(3)H]-thymidine incorporation, mRNA expression of cyclooxygenases and EP receptors by RT-PCR on cultured cells and in homogenized growth plates. To determine cellular expression, frozen sections of rat tibial growth plate and primary chondrocyte cultures were stained using immunohistochemistry with polyclonal antibodies directed towards COX-1, COX-2, EP1, EP2, EP3, and EP4. Cultured growth plate chondrocytes transiently secreted PGE(2 )into the culture medium. Although both enzymes were expressed in chondrocytes in vitro and in vivo, it appears that mainly COX-2 contributed to PGE(2)-dependent proliferation. Exogenously added PGE(2 )stimulated DNA synthesis in a dose-dependent fashion and gave a bell-shaped curve with a maximum at 10(-8 )M. The EP1/EP3 specific agonist sulprostone and the EP1-selective agonist ONO-D1-004 increased DNA synthesis. The effect of PGE(2 )was suppressed by ONO-8711. The expression of EP1, EP2, EP3, and EP4 receptors in situ and in vitro was observed; EP2 was homogenously expressed in all zones of the growth plate in situ, whereas EP1 expression was inhomogenous, with spared cells in the reserve zone. In cultured cells these four receptors were expressed in a subset of cells only. The most intense staining for the EP1 receptor was found in polygonal cells surrounded by matrix. Expression of receptor protein for EP3 and EP4 was observed also in rat growth plates. In cultured chrondrocytes, however, only weak expression of EP3 and EP4 receptor was detected. We suggest that in growth plate chondrocytes, COX-2 is responsible for PGE(2 )release, which stimulates cell proliferation via the EP1 receptor