Pro-inflammatory Enzyme Cyclooxygenase-2 Research Articles

Abstract 4118: To dine with red wine: Systems approach to studying resveratrol's protection against inflammatory colorectal cancer

Abstract The relationship between inflammation and the development of colorectal cancer has received considerable attention over the last decade. Novel agents with anti-inflammatory properties appear to have beneficial effects in reducing the risk of developing colorectal cancer. Resveratrol, a naturally occurring polyphenolic compound, has been studied extensively with considerable amounts of evidence demonstrating its efficacy as an anti-inflammatory and anti-cancer agent. Given the number and diversity of the reported protective roles of resveratrol, we used an unbiased systems approach to elucidate the underlying mechanisms of resveratrol's protection against colitis-induced colorectal cancer. Utilizing genomics, metabolomics and microbiome analysis in a pre-clinical model of colitis-induced colorectal carcinogenesis, we have linked the anti-inflammatory actions of resveratrol to both known targets and other novel potential molecular mechanisms. In a study, we treated mice with chemical carcinogen azoxymethane (AOM) and then exposed the mice to colonic irritant dextran sodium sulfate (DSS) to induce colitis-associated colon carcinogenesis. We found that a diet enriched in resveratrol (300ppm) prevented polyp formation, as well as, prevented polyps from progressing to adenomacarcinomas. Resveratrol treatment promoted the early recovery of DSS-induced inflammation with less severity of leukocyte infiltration and lower levels of IL-6, IL-8, and IL-10. Interestingly, the level of IL-22, which contributes to host defense at mucosal surfaces, was also significantly reduced with resveratrol. The metabolomic profile of DSS-treated control animals showed an elevation of microbial derived metabolites in the serum, which was ameliorated with resveratrol treatment. Together, resveratrol appears to preserve the intestinal barrier during early inflammation. Prior studies have shown resveratrol's role in inhibiting several key enzymes active during tumor progression. We have correlated such findings with metabolomic biomarkers. We found a significant reduction in arachidonic acid, the limiting reagent of pro-inflammatory enzyme Cyclooxygenase-2 (COX-2). We also saw a reduction in spermidine. This polyamine is a product of Ornithine decarboxylase (ODC) activity that promotes proliferation. We have linked some of these metabolomic changes to changes in gene expression. These pre-clinical studies are consistent with that of previous studies and demonstrate the efficacy of resveratrol in preventing colitis associated colorectal cancer. A systems approach to study the chemopreventive efficacy of anti-inflammatory compounds like resveratrol may be an important paradigm for discovering the chemopreventive mechanisms of these agents. Citation Format: Shakir M. Saud, Amiran Dzutsev, Giorgio Trinchieri, Nancy Colburn, Matthew R. Young, Young Kim. To dine with red wine: Systems approach to studying resveratrol's protection against inflammatory colorectal cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4118. doi:10.1158/1538-7445.AM2014-4118

The cardiovascular effect of the uremic solute indole-3 acetic acid.

In CKD, uremic solutes may induce endothelial dysfunction, inflammation, and oxidative stress, leading to increased cardiovascular risk. We investigated whether the uremic solute indole-3 acetic acid (IAA) predicts clinical outcomes in patients with CKD and has prooxidant and proinflammatory effects. We studied 120 patients with CKD. During the median study period of 966 days, 29 patients died and 35 experienced a major cardiovascular event. Kaplan-Meier analysis revealed that mortality and cardiovascular events were significantly higher in the higher IAA group (IAA>3.73 µM) than in the lower IAA group (IAA<3.73 µM). Multivariate Cox regression analysis demonstrated that serum IAA was a significant predictor of mortality and cardiovascular events after adjustments for age and sex; cholesterol, systolic BP, and smoking; C-reactive protein, phosphate, body mass index, and albumin; diastolic BP and history of cardiovascular disease; and uremic toxins p-cresyl sulfate and indoxyl sulfate. Notably, IAA level remained predictive of mortality when adjusted for CKD stage. IAA levels were positively correlated with markers of inflammation and oxidative stress: C-reactive protein and malondialdehyde, respectively. In cultured human endothelial cells, IAA activated an inflammatory nongenomic aryl hydrocarbon receptor (AhR)/p38MAPK/NF-κB pathway that induced the proinflammatory enzyme cyclooxygenase-2. Additionally, IAA increased production of endothelial reactive oxygen species. In conclusion, serum IAA may be an independent predictor of mortality and cardiovascular events in patients with CKD. In vitro, IAA induces endothelial inflammation and oxidative stress and activates an inflammatory AhR/p38MAPK/NF-κB pathway.

Underwater trauma causes a long-term specific increase in the expression of cyclooxygenase-2 in the ventral CA1 of the hippocampus

The pro-inflammatory enzyme cyclooxygenase-2 (COX-2) is regularly expressed in the hippocampal neurons, but its role in emotional trauma is not known. Here we show that a single acute stress caused by a near-drowning experience results in heightened anxiety-like behavior one month after the trauma. Biochemical analyses of dorsal and ventral hippocampal CA1, CA3 and dentate gyrus revealed decreased ubiquitination and elevated levels of COX-2 in the traumatized animals only in the ventral CA1. To reveal the identity of the ubiquitin E3 ligase that targets COX-2, we tested the effect of several representative E3 ligases on COX-2 expression in vitro. We found that while AIP4 and Nedd4 had no effect, Mdm2 lowered COX-2 expression by nearly 50%, an effect that was not observed by its dominant negative form. To test whether this also occurs in the hippocampus, we immunoprecipitated Mdm2 from dorsal and ventral CA1 of traumatized and control animals and probed for the presence of COX-2. Our results showed that the levels of Mdm2 were not affected by the trauma but there was significantly less COX-2 associated with Mdm2 in the ventral but not dorsal CA1 of the traumatized animals. Together these data propose that an increase in COX-2 expression in ventral CA1 following trauma is likely due to its attenuated degradation. Unraveling the pathways and mechanisms that control hippocampal COX-2 degradation is important to boost the development of novel therapeutic approaches designed to treat stress-related pathologies.

AETIQ: A Novel Synthetic Compound with Anti-inflammatory Properties in Activated Microglia

Neuroinflammation is believed to be involved in the pathogenesis of neurodegenerative diseases. Our novel synthetic compound 2-acetyl-1-ethyl-7-hydroxy-6-methoxy-1, 2, 3, 4-tetrahydroisoquinoline (AETIQ) was tested for its anti-inflammatory properties in activated microglial BV-2 cells. AETIQ attenuated nitric oxide (NO) and reactive oxygen species generation. It also downregulated the production of the proinflammatory enzymes inducible NO synthase, cyclooxygenase-2, and matrix metalloproteinase-3 at both mRNA and protein levels. Furthermore, AETIQ suppressed generation of the proinflammatory cytokines IL-1β and TNF-α as determined by reverse transcriptase-polymerase chain reaction (RT-PCR) and ELISA assays. AETIQ attenuated NFκB signaling by downregulating NFκB nuclear translocation. The compound was stable against the liver enzymes in the microsomal and S9 fractions, indicative of good bioavailability. These results suggested that AETIQ might be utilized towards development of a therapy for neuroinflammation-related diseases.

Amomum tsao-ko suppresses lipopolysaccharide-induced inflammatory responses in RAW264.7 macrophages via Nrf2-dependent heme oxygenase-1 expression.

Amomum tsao-ko Crevost et Lemaire, used as a spice in Asia, is an important source of Chinese cuisine and traditional Chinese medicines. A. tsao-ko is reported to exert a variety of biological and pharmacological activities, including anti-proliferative, anti-oxidative and neuroprotective effects. In this study, NNMBS227, consisting of the ethanol extract of A. tsao-ko, exhibited potent anti-inflammatory activities in RAW264.7 macrophages. We investigated the effect of NNMBS227 in the suppression of pro-inflammatory mediators, including pro-inflammatory enzymes (inducible nitric oxide synthase and cyclooxygenase-2) and cytokines (tumor necrosis factor-α and interleukin-1β) in LPS stimulated macrophages. NNMBS227 also inhibited the phosphorylation and degradation of IκB-α, as well as the nuclear translocation of nuclear factor kappa B (NF-κB) p65 caused by stimulation with LPS. In addition, NNMBS227 induced heme oxygenase (HO)-1 expression through the nuclear translocation of nuclear factor E2-related factor 2 (Nrf2) in macrophages. Using tin protoporphyrin (SnPP), an HO activity inhibitor, we confirmed an association between the anti-inflammatory effects of NNMBS227 and the up-regulation of HO-1. These findings suggest that Nrf2-dependent increases in the expression of HO-1 induced by NNMBS227 conferred anti-inflammatory activities in LPS stimulated RAW264.7 macrophages.

Pomegranate polyphenolics suppressed azoxymethane-induced colorectal aberrant crypt foci and inflammation: possible role of miR-126/VCAM-1 and miR-126/PI3K/AKT/mTOR

The antitumorigenic activities of polyphenols such as ellagitannins and anthocyanins in pomegranate (Punica granatum L.) have been previously studied where cytotoxic, anti-inflammatory and antioxidant effects were evident in various cancer models. The objective of this study was to investigate the role of miR-126/vascular cell adhesion molecule 1 (VCAM-1) and miR-126/phosphatidylinositol-3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) in pomegranate-mediated anti-inflammatory and anticarcinogenic effects in vivo and in vitro. Sprague-Dawley rats (n = 10 per group) received pomegranate juice (2504.74 mg gallic acid equivalents/l) or a polyphenol-free control beverage ad libitum for 10 weeks and were injected with azoxymethane (AOM) subcutaneously (15mg/kg) at weeks 2 and 3. Consumption of pomegranate juice suppressed the number of aberrant crypt foci (ACF) and dysplastic ACF by 29 and 53.5% (P = 0.05 and 0.04), respectively, and significantly lowered proliferation of mucosa cells. Pomegranate juice significantly downregulated proinflammatory enzymes nitric oxide synthase and cyclooxygenase-2 messenger RNA (mRNA) and protein expression. In addition, it suppressed nuclear factor-κB and VCAM-1 mRNA and protein expression in AOM-treated rats. Pomegranate also inhibited phosphorylation of PI3K/AKT and mTOR expression and increased the expression of miR-126. The specific target and functions of miR-126 were investigated in HT-29 colon cancer cell lines. In vitro, the involvement of miR-126 was confirmed using the antagomiR for miR-126, where pomegranate reversed the effects of the antagomiR on the expression of miR-126, VCAM-1 and PI3K p85β. In summary, therapeutic potentials of pomegranate in colon tumorigenesis were due in part to targeting miR-126-regulated pathways, which contributes in the underlying anti-inflammatory mechanisms.

Anesthetic Effects on γ-Aminobutyric Acid A Receptors

Anesthetic Effects on γ-Aminobutyric Acid A Receptors

Angiotensin II activates NF-κB through AT1A receptor recruitment of β-arrestin in cultured rat vascular smooth muscle cells

Activation of the angiotensin type 1A receptor (AT1AR) in rat aorta vascular smooth muscle cells (RASMC) results in increased synthesis of the proinflammatory enzyme cyclooxygenase-2 (COX-2). We previously showed that nuclear localization of internalized AT1AR results in activation of transcription of the gene for COX-2, i.e., prostaglandin-endoperoxide synthase-2. Others have suggested that ANG II stimulation of COX-2 protein synthesis is mediated by NF-κB. The purpose of the present study was to examine the interrelationship between AT1AR activation, β-arrestin recruitment, and NF-κB activation in the ability of ANG II to increase COX-2 protein synthesis in RASMC. In the present study we utilized RASMC, inhibitors of the NF-κB pathway, β-arrestin knockdown, radioligand binding, immunoblotting, and immunofluorescence to characterize the roles of AT1AR internalization, NF-κB activation, and β-arrestin in ANG II-induced COX-2 synthesis. Ro-106-9920 or parthenolide, agents that inhibit the initial steps of NF-κB activation, blocked ANG II-induced p65 NF-κB nuclear localization, COX-2 protein expression, β-arrestin recruitment, and AT1AR internalization without inhibiting ANG II-induced p42/44 ERK activation. Curcumin, an inhibitor of NF-κB-induced transcription, blocked ANG II-induced COX-2 protein expression without altering AT1AR internalization, ANG II-induced p65 NF-κB nuclear localization, or p42/44 ERK activation. Small interfering RNA-induced knockdown of β-arrestin-1 and -2 inhibited ANG II-induced p65 NF-κB nuclear localization. In vascular smooth muscle cells, internalization of the activated AT1AR mediated by β-arrestins activates the NF-κB pathway, producing nuclear localization of the transcription factor and initiation of COX-2 protein synthesis, thereby linking internalization of the receptor with the NF-κB pathway.

Evaluation of the immunoexpression of proinflammatory enzyme COX-2 and P53 gene in ulcerative colitis-associated dysplasia

Background and aim Ulcerative colitis (UC) patients are at increased risk for colorectal dysplasia and colorectal cancer. The aim of this study was to analyze the expression alteration of proinflammatory enzyme cyclooxygenase-2 (COX-2) and the tumor-suppressor gene P53 to clarify whether these alterations may play a role in the pathogenesis of dysplasia, which is a precancerous lesion. Patients and methods This study was conducted on 40 individuals who were divided into three groups: 15 UC patients without dysplasia, 15 UC patients with dysplasia, and 10 participants with normal colonoscopy and histopathology as a control group. Results There was a significant difference in the expression of Cox-2 and p53 in UC-related dysplasia compared with the UC group without dysplasia. There was a correlation between COX-2 and P53 expression and severity and duration of disease. Conclusion Addition of immunohistochemical analysis of Cox-2 enzyme and p53 gene to routine histologic assessment may improve the accuracy of early detection of dysplasia, which is a precancerous lesion, and thus decrease mortality from colorectal cancer.

Possible involvement of galectin-3 in microglial activation in the hippocampus with trimethyltin treatment

Trimethyltin (TMT) is an organotin neurotoxicant with effects that are selectively localized to the limbic system (especially the hippocampus), which produces memory deficits and temporal lobe seizures. Galectin-3 (Gal-3) is a beta-galactoside-binding lectin that is important in cell proliferation and regulation of apoptosis. The present study evaluated the temporal expression of Gal-3 in the hippocampus of adult BALB/c mice after TMT treatment (i.p., 2.5mg/kg). Western blotting analyses showed that Gal-3 immunoreactivity began to increase 2days after treatment; the immunoreactivity peaked significantly within 4days after treatment but significantly declined between days 4 and 8. Immunohistochemical analysis indicated that Gal-3 expression was very rare in the hippocampi of vehicle-treated controls. However, Gal-3 immunoreactivity appeared between 2 and 8days after TMT treatment and was primarily localized to the hippocampal dentate gyrus (DG), in which neuronal degeneration occurred. The immunoreactivity was detected predominantly in most of the Iba1-positive microglia and in some GFAP-positive astrocytes of the hippocampal DG. Furthermore, Gal-3 expression co-localized with the pro-inflammatory enzymes cyclooxygenase-2 and inducible nitric oxide synthase in the hippocampal DG. Therefore, we suggest that Gal-3 is involved in the inflammatory process of neurodegenerative disorder induced by organotin intoxication.

Calpain inhibition attenuated morphological and molecular changes in skeletal muscle of experimental allergic encephalomyelitis rats

Muscle weakness and atrophy are important manifestations of multiple sclerosis (MS). To investigate the pathophysiological mechanisms of skeletal muscle change in MS, we induced experimental autoimmune encephalomyelitis (EAE) in Lewis male rats and examined morphological and molecular changes in skeletal muscle. We also treated EAE rats with calpepetin, a calpain inhibitor, to examine its beneficial effects on skeletal muscle damage. Morphological changes in muscle tissue of EAE rats included smaller and irregularly shaped muscle fibers and fibrosis. Western blot analysis demonstrated increased calpain:calpastatin ratio, inflammation-related transcription factors (nuclear factor-κB:inhibitor of κB α ratio), and proinflammatory enzymes (cyclooxygenase-2). TUNEL-positive myonuclei in skeletal muscle cells of EAE rats indicated cell death. In addition, markers of apoptotic cell death (Bax:Bcl-2 ratio and caspase-12 protein levels) were elevated. Expression of muscle-specific ubiquitin ligases (muscle atrophy F-box and muscle ring finger protein 1), was upregulated in muscle tissue of EAE-vehicle animals. Both prophylactic and therapeutic treatment with calpeptin partially attenuated muscle changes noted in EAE animals. These results indicate that morphological and molecular changes including apoptotic cell death and protein breakdown develop in skeletal muscle of EAE animals and that these changes can be reversed by calpain inhibition.

Experience‐dependent brain plasticity after stroke: effect of ibuprofen and poststroke delay

Despite indications that brain plasticity may be enhanced after stroke, we have described impairment of experience-dependent plasticity in rat cerebral cortex neighboring the stroke-induced lesion. Photothrombotic stroke was centered behind the barrel cortex in one cerebral hemisphere of rats. Plasticity of cortical representation of one row of vibrissae was induced by sensory deprivation of all surrounding whiskers for 1 month, and visualized with [(14)C]-2-deoxyglucose autoradiography. In control rats deprivation resulted in an enlargement of functional cortical representation of the spared row of vibrissae. After a focal stroke neighbouring the barrel cortex, no plasticity of the spared row representation was found. Investigation of plastic changes with deprivation initiated 1 week and 1 month after stroke have shown that later poststroke onset of deprivation resulted in a partial recovery of cortical plasticity in the barrel field. Western blot analysis of proinflammatory enzyme cyclooxygenase-2 (COX-2) expression revealed its strong upregulation in the barrel cortex 24 h after stroke. When chronic treatment with the anti-inflammatory drug ibuprofen (10 mg/kg or 20 mg/kg) accompanied deprivation, plasticity was restored. Ibuprofen applied before the ischemia also prevented the poststroke upregulation of COX-2. The results strongly suggest that poststroke impairment of experience-dependent cortical plasticity is caused by stroke-induced inflammatory reactions that subside with poststroke delay and can be at least partially ameliorated by pharmacological treatment.

Chemotactic and mitogenic stimuli of neuronal apoptosis in patients with medically intractable temporal lobe epilepsy

To identify the upstream signals of neuronal apoptosis in patients with medically intractable temporal lobe epilepsy (TLE), we evaluated by immunohistochemistry and confocal microscopy brain tissues of 13 TLE patients and 5 control patients regarding expression of chemokines and cell-cycle proteins. The chemokine RANTES (CCR5) and other CC-chemokines and apoptotic markers (caspase-3, -8, -9) were expressed in lateral temporal cortical and hippocampal neurons of TLE patients, but not in neurons of control cases. The chemokine RANTES is usually found in cytoplasmic and extracellular locations. However, in TLE neurons, RANTES was displayed in an unusual location, the neuronal nuclei. In addition, the cell-cycle regulatory transcription factor E2F1 was found in an abnormal location in neuronal cytoplasm. The pro-inflammatory enzyme cyclooxygenase-2 and cytokine interleukin-1β were expressed both in neurons of patients suffering from temporal lobe epilepsy and from cerebral trauma. The vessels showed fibrin leakage, perivascular macrophages and expression of IL-6 on endothelial cells. In conclusion, the cytoplasmic effects of E2F1 and nuclear effects of RANTES might have novel roles in neuronal apoptosis of TLE neurons and indicate a need to develop new medical and/or surgical neuroprotective strategies against apoptotic signaling by these molecules. Both RANTES and E2F1 signaling are upstream from caspase activation, thus the antagonists of RANTES and/or E2F1 blockade might be neuroprotective for patients with medically intractable temporal lobe epilepsy. The results have implications for the development of new medical and surgical therapies based on inhibition of chemotactic and mitogenic stimuli of neuronal apoptosis in patients with medically intractable temporal lobe epilepsy.

Involvement of Heme Oxygenase-1 Induction in the Cytoprotective and Immunomodulatory Activities ofViola patriniiin Murine Hippocampal and Microglia Cells

A number of diseases that lead to injury of the central nervous system are caused by oxidative stress and inflammation in the brain. In this study, NNMBS275, consisting of the ethanol extract of Viola patrinii, showed potent antioxidative and anti-inflammatory activity in murine hippocampal HT22 cells and BV2 microglia. NNMBS275 increased cellular resistance to oxidative injury caused by glutamate-induced neurotoxicity and reactive oxygen species generation in HT22 cells. In addition, the anti-inflammatory effects of NNMBS275 were demonstrated by the suppression of proinflammatory mediators, including proinflammatory enzymes (inducible nitric oxide synthase and cyclooxygenase-2) and cytokines (tumor necrosis factor-α and interleukin-1β). Furthermore, we found that the neuroprotective and anti-inflammatory effects of NNMBS275 were linked to the upregulation of nuclear transcription factor-E2-related factor 2-dependent expression of heme oxygenase-1 in HT22 and BV2 cells. These results suggest that NNMBS275 possesses therapeutic potential against neurodegenerative diseases that are induced by oxidative stress and neuroinflammation.

Anti-Neuroinflammatory effects of the extract of Achillea fragrantissima

BackgroundThe neuroinflammatory process plays a central role in the initiation and progression of neurodegenerative diseases such as Parkinson's and Alzheimer's diseases, and involves the activation of brain microglial cells. During the neuroinflammatory process, microglial cells release proinflammatory mediators such as cytokines, matrix metalloproteinases (MMP), Reactive oxygen species (ROS) and nitric oxide (NO). In the present study, extracts from 66 different desert plants were tested for their effect on lipopolysaccharide (LPS) - induced production of NO by primary microglial cells. The extract of Achillea fragrantissima (Af), which is a desert plant that has been used for many years in traditional medicine for the treatment of various diseases, was the most efficient extract, and was further studied for additional anti-neuroinflammatory effects in these cells.MethodsIn the present study, the ethanolic extract prepared from Af was tested for its anti-inflammatory effects on lipopolysaccharide (LPS)-activated primary cultures of brain microglial cells. The levels of the proinflammatory cytokines interleukin1β (IL-1β) and tumor necrosis factor-α (TNFα) secreted by the cells were determined by reverse transcriptase-PCR and Enzyme-linked immunosorbent assay (ELISA), respectively. NO levels secreted by the activate cells were measured using Griess reagent, ROS levels were measured by 2'7'-dichlorofluorescein diacetate (DCF-DA), MMP-9 activity was measured using gel zymography, and the protein levels of the proinflammatory enzymes cyclooxygenase-2 (COX-2) and induced nitric oxide synthase (iNOS) were measured by Western blot analysis. Cell viability was assessed using Lactate dehydrogenase (LDH) activity in the media conditioned by the cells or by the crystal violet cell staining.ResultsWe have found that out of the 66 desert plants tested, the extract of Af was the most efficient extract and inhibited ~70% of the NO produced by the LPS-activated microglial cells, without affecting cell viability. In addition, this extract inhibited the LPS - elicited expression of the proinflammatory mediators IL-1β, TNFα, MMP-9, COX-2 and iNOS in these cells.ConclusionsThus, phytochemicals present in the Af extract could be beneficial in preventing/treating neurodegenerative diseases in which neuroinflammation is part of the pathophysiology.

Neuroprotective and anti-inflammatory effects of mollugin via up-regulation of heme oxygenase-1 in mouse hippocampal and microglial cells

Mollugin, a bioactive phytochemical isolated from Rubia cordifolia L. (Rubiaceae), exhibits antimutagenic activity, antitumor activity, antiviral activity, and inhibitory activity in arachidonic acid- and collagen-induced platelet aggregation. In this study, we investigated the effects of mollugin as a neuroprotective agent in glutamate-induced neurotoxicity in the mouse hippocampal HT22 cell line and as an anti-inflammatory agent in lipopolysaccharide-induced microglial activation in BV2 cells. Mollugin showed potent neuroprotective effects against glutamate-induced neurotoxicity and reactive oxygen species generation in mouse hippocampal HT22 cells. In addition, the anti-inflammatory effects of mollugin were demonstrated by the suppression of pro-inflammatory mediators, including pro-inflammatory enzymes (inducible nitric oxide synthase and cyclooxygenase-2) and cytokines (tumor necrosis factor-α and interleukin-6). Furthermore, we found that the neuroprotective and anti-inflammatory effects of mollugin were linked to the up-regulation of the expression of heme oxygenase (HO)-1 and the activity of HO in HT22 and BV2 cells. In addition, the effects of mollugin resulted in the nuclear accumulation of nuclear factor-E2-related factor 2 (Nrf2) in HT22 and BV2 cells. Furthermore, mollugin also activated the p38 mitogen-activated protein kinase (MAPK) pathway both in HT22 and BV2 cells. These results suggest that mollugin may be a promising candidate for the treatment of neurodegenerative diseases related to neuroinflammation.

Docosahexaenoic acid-derived neuroprotectin D1 induces neuronal survival via secretase- and PPARγ-mediated mechanisms in Alzheimer's disease models.

Neuroprotectin D1 (NPD1) is a stereoselective mediator derived from the omega-3 essential fatty acid docosahexaenoic acid (DHA) with potent inflammatory resolving and neuroprotective bioactivity. NPD1 reduces Aβ42 peptide release from aging human brain cells and is severely depleted in Alzheimer's disease (AD) brain. Here we further characterize the mechanism of NPD1's neurogenic actions using 3xTg-AD mouse models and human neuronal-glial (HNG) cells in primary culture, either challenged with Aβ42 oligomeric peptide, or transfected with beta amyloid precursor protein (βAPP)sw (Swedish double mutation APP695sw, K595N-M596L). We also show that NPD1 downregulates Aβ42-triggered expression of the pro-inflammatory enzyme cyclooxygenase-2 (COX-2) and of B-94 (a TNF-α-inducible pro-inflammatory element) and apoptosis in HNG cells. Moreover, NPD1 suppresses Aβ42 peptide shedding by down-regulating β-secretase-1 (BACE1) while activating the α-secretase ADAM10 and up-regulating sAPPα, thus shifting the cleavage of βAPP holoenzyme from an amyloidogenic into the non-amyloidogenic pathway. Use of the thiazolidinedione peroxisome proliferator-activated receptor gamma (PPARγ) agonist rosiglitazone, the irreversible PPARγ antagonist GW9662, and overexpressing PPARγ suggests that the NPD1-mediated down-regulation of BACE1 and Aβ42 peptide release is PPARγ-dependent. In conclusion, NPD1 bioactivity potently down regulates inflammatory signaling, amyloidogenic APP cleavage and apoptosis, underscoring the potential of this lipid mediator to rescue human brain cells in early stages of neurodegenerations.

Topical Application of Compound Ibuprofen Suppresses Pain by Inhibiting Sensory Neuron Hyperexcitability and Neuroinflammation in a Rat Model of Intervertebral Foramen Inflammation

There is lack of evidence that topical application of an anti-inflammatory reagent could reduce pain due to intervertebral foramen (IVF) inflammation (IVFI). We investigated analgesic effects and underlying mechanisms of topical application of a compound ibuprofen cream (CIC) onto the surface of back skin covering the inflamed L(5) IVF in a rat model. Repetitive CIC treatment (~.54 g each treatment daily for 5 consecutive days) significantly reduces severity and duration of IVFI-induced thermal hyperalgesia and mechanical allodynia by 80 to 100% and 50 to 66%, respectively. Electrophysiological studies and Western blot analysis demonstrated that CIC treatment significantly inhibited hyperexcitability of the inflamed dorsal root ganglion (DRG) neurons and upregulation of Nav1.7 and Nav1.8 protein, respectively. Pathological manifestations of the inflamed DRG were also markedly improved following CIC treatment. Further, in the inflamed DRGs, phosphorylation and expression of transcription factor NF-κB and pro-inflammatory enzyme cyclooxygenase-2 (COX-2) were significantly increased, while a cytokine IL-1β level was increased. IVFI-induced upregulation of these molecules was significantly inhibited by CIC treatment. This study provides evidence that an anti-inflammatory reagent can be used topically to suppress pain due to IVFI and/or DRG inflammation through inhibition of sensory neuron hyperexcitability and the immune and inflammatory responses. This study suggests a convenient and safe clinical intervention for treating pain due to intervertebral foramen inflammation and similar syndromes.

Cancer Chemopreventive Activity of Maslinic Acid: Suppression of COX-2 Expression and Inhibition of NF-κB and AP-1 Activation in Raji Cells

Chronic inflammation is one of the predisposing factors for neoplastic transformation. Targeting inflammation through suppression of the pro-inflammatory pathway by dietary phytochemicals provides an important strategy for cancer prevention. Maslinic acid is a novel natural triterpenoid known to inhibit proliferation and induce apoptosis in some tumor cell lines. Although maslinic acid has cytotoxic and pro-apoptotic effects on cancer cells, the underlying mechanisms of its effects on the inflammatory pathway have yet to be elucidated. It has been reported that abnormal expression of pro-inflammatory enzyme cyclooxygenase-2 (COX-2) causes promotion of cellular proliferation, suppression of apoptosis, enhancement of angiogenesis and invasiveness. In the present study, the suppressive effect of maslinic acid on COX-2 expression and the binding activity of upstream transcription factors NF- κB and AP-1, which are known to regulate COX-2 transcriptional activation, were assessed using Raji cells. The anti-inflammatory action of maslinic acid was benchmarked against oleanolic acid and other standard drugs. Western blot analysis and electrophoretic mobility shift assay (EMSA) were employed to analyze COX-2 expression as well as NF- κB and AP-1 binding activity. Our results showed that maslinic acid suppresses COX-2 expression in a concentration-dependent manner. Likewise, the constitutive nuclear NF- κB (p65) activity as well as phorbol 12-myristate 13-acetate (PMA)- and sodium N-butyrate (SnB)-induced AP-1 binding activity in Raji cells were significantly reduced following treatment with maslinic acid. Since maslinic acid suppresses COX-2 expression in Raji cells at concentrations that also lowered the NF- κB (p65) and AP-1 binding activity, it is possible that the suppression of COX-2 by this natural triterpenoid might be achieved, at least in part, via the NF- κB and AP-1 signaling pathways.

Anti-neuroinflammatory effects of geranium oil in microglial cells

Microglial cells are major immune cells in the brain, and their activation is involved in neurodegenerative diseases such as Alzheimer’s disease. Activated microglial cells secrete proinflammatory factors including nitric oxide (NO), which is an important mediator of neuroinflammation and neuronal cell death. In the present study, we demonstrated that geranium oil inhibited NO production, as well as the expression of the proinflammatory enzymes cyclooxygenase-2 (COX-2) and induced nitric oxide synthase (iNOS) in primary cultures of activated microglial cells. When tested at their natural relative concentrations in the oil, none of the major constituents of the oil could inhibit NO production. However, at higher concentrations, citronellol exhibited an inhibitory activity. The results suggest a possible synergistic interaction between these components. Thus, geranium oil might be beneficial in the prevention/treatment of neurodegenerative diseases where neuroinflammation is part of the pathophysiology.