Wnt/ß-Catenin Signaling Induces Integrin α4ß1 in T Cells and Promotes a Progressive Neuroinflammatory Disease in Mice.

The mechanisms leading to autoimmune and inflammatory diseases in the CNS have not been elucidated. The environmental triggers of the aberrant presence of CD4+ T cells in the CNS are not known. In this article, we report that abnormal ß-catenin expression in T cells drives a fatal neuroinflammatory disease in mice that is characterized by CNS infiltration of T cells, glial activation, and progressive loss of motor function. We show that enhanced ß-catenin expression in T cells leads to aberrant and Th1-biased T cell activation, enhanced expression of integrin α4ß1, and infiltration of activated T cells into the spinal cord, without affecting regulatory T cell function. Importantly, expression of ß-catenin in mature naive T cells was sufficient to drive integrin α4ß1 expression and CNS migration, whereas pharmacologic inhibition of integrin α4ß1 reduced the abnormal T cell presence in the CNS of ß-catenin-expressing mice. Together, these results implicate deregulation of the Wnt/ß-catenin pathway in CNS inflammation and suggest novel therapeutic strategies for neuroinflammatory disorders.

Use of Mometasone furoate in prolonged treatment of experimental spinal cord injury in mice: A comparative study of three different glucocorticoids.

Traumatic spinal cord injury (SCI) represents one of the most disabling injuries of the human body causing temporary or permanent sensory and/or motor system deficit, particularly hind limb locomotor function impairment. At present, steroidal inflammatory drugs, in particular methylprednisolone sodium succinate (MPSS) are the first line choice treatment of acute SCI. Despite progress in pharmacological, surgical and rehabilitative treatment approaches, SCI still remains a very complex medical and psychological challenge, with no curative therapy available. The aim of the present study was to compare the efficacy of MPSS in respect to other GCs such as dexamethasone (Dex) and mometasone furoate (MF) in an in vitro suitable model of LPS-induced inflammation in J774 cells as well as in an in vivo experimental mouse SCI (compression model). In both the in vitro and in vivo experiments, MF resulted surprisingly more potent than Dex and MPSS. In detail, mice sacrificed seven days after induction of SCI trauma resulted not only in tissue damage, cellular infiltration, fibrosis, astrocyte activation, iNOS expression, extracellular signal regulated kinase 1/2 phosphorylation in injured tissue, poly (ADP-ribose) polymerase 1 (PARP-1) activation but also apoptosis (Bax and Bcl-2 expression). All three GCs demonstrated the ability to modulate inflammatory, oxidative as well as apoptotic pathways, but MF demonstrated the best efficacy, while Dex and MPSS showed alternative potency with a different degree of protection. Therefore, we can conclude that MF is the best candidate for post-traumatic chronic treatment, since it ameliorates different molecular pathways involved in the damage's propagation to the surrounding areas of the injured spinal cord.

4(α-l-rhamnosyloxy)-benzyl isothiocyanate, a bioactive phytochemical that attenuates secondary damage in an experimental model of spinal cord injury.

4(α-l-Rhamnosyloxy)-benzyl isothiocyanate (glucomoringin isothiocyanate; GMG-ITC) is released from the precursor 4(α-l-rhamnosyloxy)-benzyl glucosinolate (glucomoringin; GMG) by myrosinase (ß-thioglucoside glucohydrolase; E.C. 3.2.1.147) catalyzed hydrolysis. GMG is an uncommon member of the glucosinolate group as it presents a unique characteristic consisting in a second glycosidic residue within the side chain. It is a typical glucosinolate found in large amounts in the seeds of Moringa oleifera Lam., the most widely distributed plant of the Moringaceae family. GMG was purified from seed-cake of M. oleifera and was hydrolyzed by myrosinase at neutral pH in order to form the corresponding GMG-ITC. This bioactive phytochemical can play a key role in counteracting the inflammatory response connected to the oxidative-related mechanisms as well as in the control of the neuronal cell death process, preserving spinal cord tissues after injury in mice. Spinal cord trauma was induced in mice by the application of vascular clips (force of 24g) for 1 min., via four-level T5-T8 after laminectomy. In particular, the purpose of this study was to investigate the dynamic changes occurring in the spinal cord after ip treatment with bioactive GMG-ITC produced 15 min before use from myrosinase-catalyzed hydrolysis of GMG (10mg/kg body weight+5 µl Myr mouse/day). The following parameters, such as histological damage, distribution of reticular fibers in connective tissue, nuclear factor (NF)-κB translocation and nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (IκB-α) degradation, expression of inducible Nitric Oxide Synthases (iNOS), as well as apoptosis, were evaluated. In conclusion, our results show a protective effect of bioactive GMG-ITC on the secondary damage, following spinal cord injury, through an antioxidant mechanism of neuroprotection. Therefore, the bioactive phytochemical GMG-ITC freshly produced before use by myrosinase-catalyzed hydrolysis of pure GMG, could prove to be useful in the treatment of spinal cord trauma.

Tuscan black kale sprout extract bioactivated with myrosinase: a novel natural product for neuroprotection by inflammatory and oxidative response during cerebral ischemia/reperfusion injury in rat.

BACKGROUND: Cerebral ischemia and reperfusion (CIR) is a pathological condition characterized by a first blood supply restriction to brain followed by the consequent restoration of blood flow and simultaneous reoxygenation. The aim of this study was to evaluate the neuroprotective effects of Tuscan black kale sprout extract (TBK-SE) bioactivated with myrosinase enzyme, assessing its capability to preserve blood-brain barrier (BBB), in a rat model of CIR. METHODS: CIR was induced in rats according to a classic model of carotid artery occlusion for a time period of 1 h and the reperfusion time was prolonged for seven days. RESULTS: By immunohistochemical evaluation and western blot analysis of brain and cerebellum tissues, our data have clearly shown that administration of bioactive TBK-SE is able to restore alterations of tight junction components (claudin-5 immunolocalization). Also, bioactive TBK-SE reduces some inflammatory key-markers (p-selectin, GFAP, Iba-1, ERK1/2 and TNF-α), as well as the triggering of neuronal apoptotic death pathway (data about Bax/Bcl-2 balance, p53 and cleaved-caspase 3) and the generation of radicalic species by oxidative stress (results focused on iNOS, nitrotyrosine and Nrf2). CONCLUSION: Taken together, our findings lead to believe that bioactive TBK-SE exerts pharmacological properties in protecting BBB integrity through a mechanism of action that involves a modulation of inflammatory and oxidative pathway as well into control of neuronal death.

Use of natural compounds in the management of diabetic peripheral neuropathy.

Nephropathy, retinopathy cardiomyopathy and peripheral neuropathy are all recognized as important complications in about 50% of diabetes mellitus (DM) patients, mostly related to a poor glycemic control or to an improper management of this pathology. In any case, amongst others, diabetic peripheral neuropathy (DPN) seems the leading and most painful complication usually affecting many DM patients. For this reason, this work was conceived to review the large variety of strategies adopted for management of DPN, starting from the most conventional therapies to arrive at alternative approaches. From this perspective, both the most popular pharmacological treatments used to respond to the poorly effect of common analgesics--non-steroidal anti-inflammatory drugs (NSAIDS) and opioids--understood as gabapentin vs. pregabalin clinical use, and the guidelines provided by Oriental Medicine as well as by a long list of natural compounds that many authors identify as possible therapeutic or alternative agents to replace or to combine with the existing therapies will be included. Moreover, in the effort to provide the widest panel of remedies, the most antique techniques of acupuncture and electrostimulation will be considered as alternative, which are useful approaches to take into account in any non-pharmacological strategy for DPN management.

Cannabinoids: new promising agents in the treatment of neurological diseases.

Nowadays, Cannabis sativa is considered the most extensively used narcotic. Nevertheless, this fame obscures its traditional employ in native medicine of South Africa, South America, Turkey, Egypt and in many regions of Asia as a therapeutic drug. In fact, the use of compounds containing Cannabis and their introduction in clinical practice is still controversial and strongly limited by unavoidable psychotropic effects. So, overcoming these adverse effects represents the main open question on the utilization of cannabinoids as new drugs for treatment of several pathologies. To date, therapeutic use of cannabinoid extracts is prescribed in patients with glaucoma, in the control of chemotherapy-related vomiting and nausea, for appetite stimulation in patients with anorexia-cachexia syndrome by HIV, and for the treatment of multiple sclerosis symptoms. Recently, researcher efforts are aimed to employ the therapeutic potentials of Cannabis sativa in the modulation of cannabinoid receptor activity within the central nervous system, particularly for the treatment of neurodegenerative diseases, as well as psychiatric and non-psychiatric disorders. This review evaluates the most recent available data on cannabinoids utilization in experimental and clinical studies, and highlights their beneficial effects in the prevention of the main neurological diseases and for the clinical treatment of symptoms with them correlated.

Anti-inflammatory and anti-apoptotic effects of (RS)-glucoraphanin bioactivated with myrosinase in murine sub-acute and acute MPTP-induced Parkinson's disease.

This study was focused on the possible neuroprotective role of (RS)-glucoraphanin, bioactivated with myrosinase enzyme (bioactive RS-GRA), in an experimental mouse model of Parkinson's disease (PD). RS-GRA is one of the most important glucosinolates, a thiosaccharidic compound found in Brassicaceae, notably in Tuscan black kale seeds. RS-GRA was extracted by one-step anion exchange chromatography, further purified by gel-filtration and analyzed by HPLC. Following, pure RS-GRA was characterized by (1)H and (13)C NMR spectrometry and the purity was assayed by HPLC analysis of the desulfo-derivative according to the ISO 9167-1 method. The obtained purity has been of 99%. To evaluate the possible pharmacological efficacy of bioactive RS-GRA (administrated at the dose of 10mg/kg, ip +5µl/mouse myrosinase enzyme), C57BL/6 mice were used in two different sets of experiment (in order to evaluate the neuroprotective effects in different phases of the disease), according to an acute (2 injections·40mg/kg MPTP) and a sub-acute (5 injections·20mg/kg MPTP) model of PD. Behavioural test, body weight changes measures and immunohistochemical localization of the main PD markers were performed and post-hoc analysis has shown as bioactive RS-GRA is able to reduce dopamine transporter degradation, tyrosine hydroxylase expression, IL-1ß release, as well as the triggering of neuronal apoptotic death pathway (data about Bax/Bcl-2 balance and dendrite spines loss) and the generation of radicalic species by oxidative stress (results focused on nitrotyrosine, Nrf2 and GFAP immunolocalization). These effects have been correlated with the release of neurotrophic factors, such as GAP-43, NGF and BDNF, that, probably, play a supporting role in the neuroprotective action of bioactive RS-GRA. Moreover, after PD-induction mice treated with bioactive RS-GRA are appeared more in health than animals that did not received the treatment both for phenotypic behaviour and for general condition (movement coordination, presence of tremors, nutrition). Overall, our results suggest that bioactive RS-GRA can protect neurons against the neurotoxicity involved in PD via an anti-apoptotic/anti-inflammatory action.

Antibacterial activity of glucomoringin bioactivated with myrosinase against two important pathogens affecting the health of long-term patients in hospitals.

Glucosinolates (GLs) are natural compounds present in species of the order Brassicales and precursors of bioactive isothiocyanates (ITCs). In the recent years, they have been studied mainly for their chemopreventive as well as novel chemotherapeutics properties. Among them 4-(α-L-rhamnosyloxy)benzyl glucosinolate (glucomoringin; GMG), purified from seeds of Moringa oleifera Lam., a plant belonging to the Moringaceae family, represents an uncommon member of the GL family with peculiar characteristics. This short communication reports new evidences about the properties of GMG and presents a new innovative utilization of the molecule. The bioactivation of GMG by myrosinase enzyme just before treatment, permits to maximize the power of the final product of the reaction, which is the 4-(α-L-rhamnosyloxy)benzyl isothiocyanate (GMG-ITC). We tested the antibiotic activity of this latter compound on two strains of pathogens affecting the health of patients in hospital, namely Staphylococcus aureus and Enterococcus casseliflavus, and on the yeast Candida albicans. Results show that the sensibility of S. aureus BAA-977 strain and E. casseliflavus to GMG-ITC treatment reveals an important possible application of this molecule in the clinical care of patients, more and more often resistant to traditional therapies.

MK801 attenuates secondary injury in a mouse experimental compression model of spinal cord trauma.

BACKGROUND: Glutamergic excitotoxicity has been shown to play a deleterious role in the pathophysiology of spinal cord injury (SCI). The aim of this study was to investigate the neuroprotective effect of dizocilpine maleate, MK801 (2 mg/Kg, 30 min and 6 hours after injury) in a mice model of SCI. The spinal cord trauma was induced by the application of vascular clips to the dura via a four-level T5-T8 laminectomy. RESULTS: Spinal cord injury in mice resulted in severe trauma characterized by edema, neutrophil infiltration and apoptosis. In this study we clearly demonstrated that administration of MK801 attenuated all inflammatory parameters. In fact 24 hours after injury, the degree of spinal cord inflammation and tissue injury (evaluated as histological score), infiltration of neutrophils, NF-κB activation, iNOS, cytokines levels (TNF-α and IL-1ß), neurotrophin expression were markedly reduced by MK801 treatment. Moreover, in a separate set of experiments, we have demonstrated that MK801 treatment significantly improved the recovery of locomotory function. CONCLUSIONS: Blockade of NMDA by MK801 lends support to the potential importance of NMDA antagonists as therapeutic agents in the treatment of acute spinal cord injury.

Adrenomedullin in inflammatory process associated with experimental pulmonary fibrosis.

BACKGROUND: Adrenomedullin (AM), a 52-amino acid ringed-structure peptide with C-terminal amidation, was originally isolated from human pheochromocytoma. AM are widely distributed in various tissues and acts as a local vasoactive hormone in various conditions. METHODS: In the present study, we investigated the efficacy of AM on the animal model of bleomycin (BLM)-induced lung injury. Mice were subjected to intratracheal administration of BLM and were assigned to receive AM daily by an intraperitoneal injection of 200 ngr/kg. RESULTS AND DISCUSSION: Myeloperoxidase activity, lung histology, immunohistochemical analyses for cytokines and adhesion molecules expression, inducible nitric oxide synthase (iNOS), nitrotyrosine, and poly (ADP-ribose) polymerase (PARP) were performed one week after fibrosis induction. Lung histology and transforming growth factor beta (TGF-ß) were performed 14 and 21 days after treatments. After bleomycin administration, AM-treated mice exhibited a reduced degree of lung damage and inflammation compared with BLM-treated mice, as shown by the reduction of (1) myeloperoxidase activity (MPO), (2) cytokines and adhesion molecules expression, (3) nitric oxide synthase expression, (4) the nitration of tyrosine residues, (5) poly (ADP-ribose) (PAR) formation, a product of the nuclear enzyme poly (ADP-ribose) polymerase (PARP) (6) transforming growth factor beta (TGF-ß) (7)and the degree of lung injury. CONCLUSIONS: Our results indicate that AM administration is able to prevent bleomycin induced lung injury through the down regulation of proinflammatory factors.

Effects of palmitoylethanolamide on release of mast cell peptidases and neurotrophic factors after spinal cord injury.

Spinal cord injury (SCI) has a significant impact on quality of life, expectancy, and economic burden, with considerable costs associated with primary care and loss of income. The complex pathophysiology of SCI may explain the difficulty in finding a suitable therapy for limiting neuronal injury and promoting regeneration. Although innovative medical care, advances in pharmacotherapy have been limited. The aim of the present study was to carefully investigate molecular pathways and subtypes of glial cells involved in the protective effect of PEA on inflammatory reaction associated with an experimental model of SCI. The compression model induced by applying an aneurysm clip to the spinal cord in mice is closer to the human situation, since it replicates the persistence of cord compression. Spinal cord trauma was induced in mice by the application of vascular clips to the dura via a four-level T5-T8 laminectomy. Repeated PEA administration (10 mg/kg i.p., 6 and 12 h after SCI) significantly reduced the degree of the severity of spinal cord trauma through the reduction of mast cell infiltration and activation. Moreover, PEA treatment significantly reduced the activation of microglia and astrocytes expressing cannabinoid CB(2) receptor after SCI. Importantly, the protective effect of PEA involved changes in the expression of neurotrophic factors, and in spinal cord dopaminergic function. Our results enhance our understanding about mechanisms related to the anti-inflammatory property of the PEA suggesting that this N-acylethanolamine may represent a crucial therapeutic intervention both diminishing the immune/inflammatory response and promoting the initiation of neurotrophic substance after SCI.

Liver X receptor agonist treatment regulates inflammatory response after spinal cord trauma.

Liver X receptor alpha (LXRalpha) and LXRbeta are members of the nuclear receptor superfamily of ligand-activated transcription factors. The aim of this study was to investigate the effects of T0901317, a potent LXR receptor ligand, in a mouse model of spinal cord injury (SCI). SCI was induced by the application of vascular clips (force of 24 g) to the dura via a four-level T5-T8 laminectomy in mice. Treatment with T0901317, 1 and 6 h after the SCI, significantly decreased (i) the degree of spinal cord inflammation and tissue injury (histological score); (ii) neutrophil infiltration (myeloperoxidase activity); (iii) inducible nitric oxide synthase expression; (iv) nitrotyrosine, lipid peroxidation, and poly-ADP-ribose formation; (v) pro-inflammatory cytokines expression; (vi) nuclear factor-kappa B activation; and (vii) apoptosis (terminal deoxynucleotidyltransferase-mediated UTP end labeling staining, FAS ligand, Bax, and Bcl-2 expression). Moreover, T0901317 significantly ameliorated the loss of limb function (evaluated by motor recovery score). These data suggest that LXR ligand may be useful in the treatment of inflammation associated with SCI.

Evidence for the role of peroxisome proliferator-activated receptor-beta/delta in the development of spinal cord injury.

Several lines of evidence suggest a biological role for peroxisome proliferator-activated receptor (PPAR)-beta/delta in the pathogenesis many diseases. The aim of the present study was to evaluate the contribution of PPAR-beta/delta in the secondary damage in experimental spinal cord injury (SCI) in mice. To this purpose, we used 4-[[[2-[3-fluoro-4-(trifluoromethyl)phenyl]-4-methyl-5-thiazolyl]methyl]thio]-2-methylphenoxy]acetic acid (GW0742), a high-affinity PPAR-beta/delta agonist. Spinal cord trauma was induced by the application of vascular clips (force of 24 g) to the dura via a four-level T5 to T8 laminectomy. SCI in mice resulted in severe trauma characterized by edema, neutrophil infiltration, production of inflammatory mediators, tissue damage, and apoptosis. GW0742 treatment (0.3 mg kg(-1) i.p.) 1 and 6 h after the SCI significantly reduced 1) the degree of spinal cord inflammation and tissue injury (histological score), 2) neutrophil infiltration (myeloperoxidase activity), 3) nitrotyrosine formation, 4) proinflammatory cytokines expression, 5) nuclear factor-kappaB activation, 6) inducible nitric-oxide synthase expression, and 6) apoptosis (terminal deoxynucleotidyl transferase dUTP nick-end labeling staining, FasL, Bax, and Bcl-2 expression). Moreover, GW0742 significantly ameliorated the recovery of limb function (evaluated by motor recovery score). To elucidate whether the protective effects of GW0742 are related to activation of the PPAR-beta/delta receptor, we also investigated the effect of PPAR-beta/delta antagonist methyl 3-({[2-(methoxy)-4 phenyl]amino}sulfonyl)-2-thiophenecarboxylate (GSK0660) on the protective effects of GW0742. GSK0660 (1 mg/kg i.p. 30 min before treatment with GW0742) significantly blocked the effect of the PPAR-beta/delta agonist and thus abolished the protective effect. Our results clearly demonstrate that GW0742 treatment reduces the development of inflammation and tissue injury associated with spinal cord trauma.

Effects of Liver x receptor agonist treatment on signal transduction pathways in acute lung inflammation.

BACKGROUND: Liver x receptor alpha (LXRalpha) and beta (LXRbeta) are members of the nuclear receptor super family of ligand-activated transcription factors, a super family which includes the perhaps better known glucocorticoid receptor, estrogen receptor, thyroid receptor, and peroxisome proliferator-activated receptors. There is limited evidence that LXL activation may reduces acute lung inflammation. The aim of this study was to investigate the effects of T0901317, a potent LXR receptor ligand, in a mouse model of carrageenan-induced pleurisy. METHODS: Injection of carrageenan into the pleural cavity of mice elicited an acute inflammatory response characterized by: accumulation of fluid containing a large number of neutrophils (PMNs) in the pleural cavity, infiltration of PMNs in lung tissues and subsequent lipid peroxidation, and increased production of nitrite/nitrate (NOx), tumor necrosis factor-alpha, (TNF-alpha) and interleukin-1beta (IL-1beta). Furthermore, carrageenan induced the expression of iNOS, nitrotyrosine and PARP, as well as induced apoptosis (TUNEL staining and Bax and Bcl-2 expression) in the lung tissues. RESULTS: Administration of T0901317, 30 min after the challenge with carrageenan, caused a significant reduction in a dose dependent manner of all the parameters of inflammation measured. CONCLUSIONS: Thus, based on these findings we propose that LXR ligand such as T0901317, may be useful in the treatment of various inflammatory diseases.

PD98059, a specific MAP kinase inhibitor, attenuates multiple organ dysfunction syndrome/failure (MODS) induced by zymosan in mice.

PD98059 (MEK1 Inhibitor) has been shown to act in vivo as a highly selective inhibitor of MEK1 activation and the MAP kinase cascade. In the present study, we have investigated the effects of PD98059, on the development of non-septic shock caused by zymosan in mice. Mice received either intraperitoneally zymosan (500mg/kg, administered i.p. as a suspension in saline) or vehicle (0.25ml/mouse saline). PD98059 (10mg/kg) was administered 1 and 6h after zymosan administration i.p. Organ failure and systemic inflammation in mice was assessed 18h after administration of zymosan and/or PD98059. Treatment of mice with PD98059 attenuated the peritoneal exudation and the migration of polymorphonuclear cells caused by zymosan. PD98059 also attenuated the lung, liver and pancreatic injury and renal dysfunction caused by zymosan as well as the increase of TNF-alpha and IL-1beta plasma levels caused by zymosan. Immunohistochemical analysis for inducible nitric oxide synthase (iNOS), nitrotyrosine, poly(ADP-ribose) (PAR), ICAM-1, P-selectin, Bax, Bcl-2 and FAS-ligand revealed positive staining in pancreatic and intestinal tissue obtained from zymosan-injected mice. The degree of staining for nitrotyrosine, iNOS, PAR, ICAM-1, P-selectin, Bax, Bcl-2 and FAS-ligand were markedly reduced in tissue sections obtained from zymosan-injected mice, which had received PD98059. Moreover treatment of mice with PD98059 (10mg/kg) attenuated the NF-kappaB activation and mitogen-activated protein kinases (MAPK) expression induced by zymosan injection. In addition, administration of zymosan caused a severe illness in the mice characterized by a systemic toxicity, significant loss of body weight and a 60% of mortality at the end of observation period. Treatment with PD98059 significantly reduced the development of systemic toxicity, the loss in body weight and the mortality (20%) caused by zymosan. This study provides evidence that PD98059 attenuates the degree of zymosan-induced non-septic shock in mice.

Effects of Hypericum Perforatum, in a rodent model of periodontitis.

BACKGROUND: Hypericum perforatum is a medicinal plant species containing many polyphenolic compounds, namely flavonoids and phenolic acids. In this study we evaluate the effect of Hypericum perforatum in animal model of periodontitis. METHODS: Periodontitis was induced in adult male Sprague-Dawley rats by placing a nylon thread ligature around the lower 1st molars. Hypericum perforatum was administered at the dose of 2 mg/kg os, daily for eight days. At day 8, the gingivomucosal tissue encircling the mandibular first molar was removed. RESULTS: Periodontitis in rats resulted in an inflammatory process characterized by edema, neutrophil infiltration and cytokine production that was followed by the recruitment of other inflammatory cells, production of a range of inflammatory mediators such as NF-κB and iNOS expression, the nitration of tyrosine residues and activation of the nuclear enzyme poly (ADP-ribose) polymerase; apoptosis and the degree of gingivomucosal tissues injury. We report here that Hypericum perforatum exerts potent anti-inflammatory effects significantly reducing all of the parameters of inflammation as described above. CONCLUSIONS: Taken together, our results clearly demonstrate that treatment with Hypericum reduces the development of inflammation and tissue injury, events associated with periodontitis.

Role of peroxisome proliferator-activated receptor-alpha in ileum tight junction alteration in mouse model of restraint stress.

Restraint stress induces permeability changes in the small intestine, but little is known about the role of endogenous peroxisome proliferator-activated receptor-alpha (PPAR-alpha) ligand in the defects of the tight junction function. In the present study, we used PPAR-alpha knockout mice to understand the roles of endogenous PPAR-alpha on ileum altered permeability function in models of immobilization stress. The absence of a functional PPAR-alpha gene in PPAR-alpha knockout mice resulted in a significant augmentation of the degree of 1) TNF-alpha production in ileum tissues; 2) the alteration of zonula occludens-1, occludin, and beta-catenin (immunohistochemistry); and 3) apoptosis (terminal deoxynucleotidyltransferase-mediated dUTP nick-end labeling staining, Bax, Bcl-2 expression). Taken together, our results demonstrate that endogenous PPAR-alpha ligands reduce the degree of tight junction permeability in the ileum tissues associated with immobilization stress, suggesting a possible role of endogenous PPAR-alpha ligands on ileum barrier dysfunction.

Effects of genetic and pharmacological inhibition of TNF-alpha in the regulation of inflammation in macrophages.

Tumour necrosis factor-alpha (TNF-alpha) is an inflammatory cytokine produced by circulating monocytes and resident macrophages during acute inflammation and is responsible for a diverse range of signalling events within cells, leading to necrosis or apoptosis. The biologic activities of TNF are mediated by two receptors TNFR1 and TNFR2, although a lot of studies demonstrated that most of the biological activities of TNF-alpha are mediated through TNFR1. In the present study, we want to evaluate the role of TNF-alpha on regulation of in vitro models of inflammation. In particular we used peritoneal macrophages, from TNF-alphaR1 knock out and TNF-alphaR1 wild-type mice, stimulated with LPS 10 microg/ml and IFN-gamma 100 U/ml. Our results showed that the deletion of TNF-alphaR1 gene significantly reduced the degree of (i) MAPK activation, (ii) IkappaB-alpha degradation, (iii) phosphorylation of Ser536 on the NF-kappaB subunit p65 and (iv) iNOS and COX-2 expression. In addition, to confirm the pivotal role of TNF-alpha on regulation of peritoneal macrophages inflammation, we have also investigated the protective effects of infliximab, a TNF-alpha chimeric mouse/human IgG1 antibody against TNF-alpha. As shown in the present study, the cell incubation with infliximab (0.1 microg/ml, 1 microg/ml and 10 microg/ml) significantly leads to a concentration-dependent inhibition of the inflammatory mediators above described. In conclusion, our study demonstrates that pharmacological and genetic inhibition of the TNF/TNFR1 binding reduce the degree of macrophages inflammation caused by LPS/IFN-gamma stimulation.

PPAR-alpha modulate the anti-inflammatory effect of glucocorticoids in the secondary damage in experimental spinal cord trauma.

Glucocorticoids (GCs) are effective anti-inflammatory agents widely used in therapeutic approach to treatment of spinal cord trauma. Previous results suggest that peroxisome proliferator activated receptor alpha (PPAR-alpha), an intracellular transcription factor activated by fatty acids, plays a role in control of secondary inflammatory process associated with spinal cord injury (SCI). With the aim to characterize the role of PPAR-alpha in GC-mediated anti-inflammatory activity, we tested the efficacy of dexamethasone (DEX), a synthetic GC specific for glucocorticoid receptor (GR), in an experimental model of spinal cord trauma induced in mice by the application of vascular clips (force of 24 g) to the dura via a four-level T5-T8 laminectomy, and comparing mice lacking PPAR-alpha (PPAR-alphaKO) with wild type (WT) mice. Results indicate that DEX-mediated anti-inflammatory activity is weakened in PPAR-alphaKO mice, as compared to WT controls. In particular, DEX was less effective in PPAR-alphaKO, compared to WT mice, as evaluated by inhibition of the degree of spinal cord inflammation and tissue injury, neutrophil infiltration, nitrotyrosine formation, pro-inflammatory cytokine expression, NF-kappaB activation, inducible nitric-oxide synthase (iNOS) expression; and apoptosis. This study indicates that PPAR-alpha can contribute to the anti-inflammatory activity of GCs in SCI.

Protective effect of melatonin against the inflammatory response elicited by crude venom from isolated nematocysts of Pelagia noctiluca (Cnidaria, Scyphozoa).

Melatonin (N-acetyl-5-methoxytryptamine) is an efficient free radical scavenger and antioxidant, both in vitro and in vivo. The role of melatonin as an immunomodulator is, in some cases, contradictory. In this study we have investigated the therapeutic efficacy of melatonin in rats subjected to Pelagia noctiluca crude venom (of the familia Pelaguiidae; and genus Pelagia) induced acute paw inflammation. In particular, injection of the venom into the paw of rats elicited an acute inflammatory response characterized by accumulation of fluid containing a large number of polymorphonuclear neutrophils in the paw and subsequent lipid peroxidation. Furthermore, the venom promoted an expression of iNOS, nitrotyrosine and the activation of the nuclear enzyme poly (ADP-ribose) polymerase as determined by immunohistochemical analysis of paw tissues. Administration of melatonin 30 min, 1 and 6 hr after the challenge with the venom, caused a significant reduction in all the parameters of inflammation measured. Thus, based on these findings we propose that melatonin may be useful a treatment of local acute inflammation induced by P. noctiluca crude venom.