Dietary eicosapentaenoic acid normalizes hippocampal omega-3 and 6 polyunsaturated fatty acid profile, attenuates glial activation and regulates BDNF function in a rodent model of neuroinflammation induced by central interleukin-1ß administration.

PURPOSE: Interleukin (IL)-1ß can activate glial cells to trigger neuroinflammation and neurodegeneration. Lower omega (n)-3 polyunsaturated fatty acids (PUFAs) and lower n-3/n-6 PUFA ratios occur in the brain of patients with Alzheimer's disease (AD). We have previously reported that an n-3 PUFA, eicosapentaenoic acid (EPA), can improve memory and attenuate neurodegeneration-like changes in animal models of AD. However, whether and how EPA modulates glial cell activity and functions remains unclear. The aim of this study was to test the hypothesis that EPA may attenuate neuroinflammation by inhibiting microglial activation and microglia-produced proinflammatory cytokines, and by enhancing the expression of astrocytes-produced neurotrophins and their receptors. METHODS: Male Long-Evans rats were fed either palm oil supplemented diet or EPA supplemented diet for 42 days. On day 36 of diet feeding, rats received an intracerebroventricular injection of IL-1ß or saline for 7 days. The glial activation, the expression of amyloid precursor protein (APP), calcium-dependent phospholipase (cPL) A2, brain-derived neurotrophic factor (BDNF) and its receptor, and PUFA profile in the hippocampus were analyzed. RESULTS: IL-1ß elevated biomarkers of microglial CD11b and astrocyte GFAP expression, increased the expression of APP, tumor-necrosis factor (TNF)-α, but reduced BDNF and its receptor (TrKB). IL-1ß also lowered n-3 EPA and docosapentaenoic acid concentrations but increased n-6 PUFAs and cPLA2 activity in the hippocampus. EPA supplement normalized the n-3 and n-6 PUFA profiles and cPLA2 levels, inhibited glial activation, reduced APP and TNF-α expression, as well as up-regulated BDNF and TrKB. CONCLUSION: Supplementation with EPA appear to have potential effects on improving glial over-activation, n3/n6 imbalance and BDNF down-regulation, which contribute to anti-inflammatory and may provide beneficial effects on inflammation-associated disease such as AD.

Modulation of MMP-2 and -9 secretion by cytokines, inducers and inhibitors in human melanoma A-2058 cells.

Melanoma, an extremely aggressive cancer, causes the most skin cancer-related deaths, due to metastasis to other areas of the body, such as lymph nodes, lungs, liver, brain or bone. It is characterized by high levels of matrix metalloproteinase (MMP)-2 and -9 secretions that degrade the extracellular matrix and basement membrane, allowing cancer cells to spread to distal organs. Various cytokines, mitogens, growth factors, inducers and inhibitors control MMP activities. We investigated the roles of these in regulation of MMP-2 and -9 in human melanoma A-2058 cells. Human A-2058 cells were grown in DMEM supplemented with 15% FBS and antibiotics in 24-well tissue culture plates. At near confluence, the cells were washed with PBS and incubated in serum-free media with phorbol 12-myristate 13-acetate (PMA) at 10, 25, 50 and 100 ng/ml; TNF-α and IL-1ß at 0.1, 1, 10 and 25 ng/ml; LPS at 10, 25, 50 and 100 µg/ml; epigallocatechin gallate (EGCG) and doxycycline (Dox) at 10, 25, 50 and 100 µM without and with PMA; a nutrient mixture (NM) containing lysine, proline, ascorbic acid and green tea extract without and with PMA at 10, 50, 100, 500 and 1,000 µg/ml; actinomycin D and cyclohexamide at 2 and 4 µM; retinoic acid and dexamethasone at 50 µM. After 24 h the media were removed and analyzed for MMP-2 and MMP-9 by zymography and densitometry. Melanoma A-2058 demonstrated strong expression of MMP-2 and slight expression of MMP-9. PMA at 100 ng/ml showed no effect on MMP-2 secretion but potently upregulated MMP-9 secretion to 400% that of control. TNF-α showed no significant overall effect on expression of MMP-2 but potent dose-dependent increased MMP-9 secretion with 200% of control at 25 ng/ml. IL-1ß showed no significant effect on MMP-2 or MMP-9 secretion by A-2058 cells, except at 25 ng/ml where MMP-2 level was reduced by ~40% and MMP-9 secretion ~50%. LPS treatment showed no significant effect on MMP-2 secretion and enhanced MMP-9 secretion up to 25 µg/ml followed by decreased level. EGCG, NM and doxycycline, without and with PMA, downregulated the expression of MMP-2 and MMP-9 in a dose-dependent manner. Actinomycin D, cyclohexamide and retinoic acid had inhibitory effects on MMP-2, while dexamethasone showed slight stimulatory effect on MMP-2 secretion. Our results showed that select cytokines, mitogens and inhibitors modulated A-2058 MMP-2 and MMP-9 expression. They suggest the clinical potential of MMP inhibitors, especially the non-toxic ones, such as the nutrient mixture and its component EGCG in management of melanoma.

Divergent effects of brain interleukin-1ß in mediating fever, lethargy, anorexia and conditioned fear memory.

The influence of brain interleukin-1 (IL-1ß) on memory processes includes both detrimental and beneficial effects. To further explore the dynamics of brain IL-1ß in mediating learning and memory during acute sickness, we injected species-homologous rat IL-1ß (100ng/5µl) or vehicle (0.1% bovine serum albumin, 5µl) directly into the cisterna magna (i.c.m.) of male Sprague-Dawley rats. We measured, in parallel, body temperature, food intake, body mass, cage activity, as well as learning and memory using contextual fear conditioning. To investigate the effects of IL-1ß on learning and memory processes we used: (1) a retrograde experiment that involved injecting rats i.c.m. with IL-1ß immediately after training in the novel context, and (2) an anterograde experiment that involved injecting rats i.c.m. with IL-1ß two hours before training in the novel context. In addition, hypothalamic and hippocampal concentrations of IL-1ß were measured at several time points following injection. Administration of IL-1ß induced fever, lethargy and anorexia for∼two-to-three days and increased the concentration of IL-1ß in the hippocampus and hypothalamus for at least eight hours. Training in the context immediately before IL-1ß administration (retrograde experiment), did not impair contextual and auditory fear memory. However, when training in the context occurred concurrently with elevated hippocampal IL-1ß levels, two hours after IL-1ß administration (anterograde experiment), contextual, but not auditory, fear memory was impaired. Our results show that there are instances where memory consolidation can occur concurrently with elevated levels of IL-1ß in the hippocampus, fever, anorexia and lethargy during acute short-term sickness.

Protective Effects of Garlic-Derived S-Allylmercaptocysteine on IL-1ß-Stimulated Chondrocytes by Regulation of MMPs/TIMP-1 Ratio and Type II Collagen Expression via Suppression of NF-κB Pathway.

BACKGROUND: Garlic-derived S-allylmercaptocysteine (SAMC) has widely been used in many disease therapies. However, the potential effects and mechanism of SAMC on IL-1ß-stimulated chondrocytes are unclear. METHODS: Chondrocytes were isolated, and 5 ng/mL of IL-1ß was added to mimic the in vitro osteoarthritis (OA) model. SAMC (20 and 60 µM) was used for the treatment in OA model. Cell viability was assessed by MTT method. Western blotting, Quantitative RT-PCR, and ELISA were performed to evaluate the mechanisms in SAMC treated OA model. RESULTS: Following 48 h of IL-1ß exposure, SAMC exhibited protection effect on IL-1ß-injured chondrocyte viability. Type II collagen was elevated with reduced degradation products, as a consequence of altered MMPs/TIMP-1 ratio after SAMC treatment in IL-1ß-treated chondrocytes. The protein and mRNA level of TNF-α in cellular supernatant and cells were downregulated in a dose-dependent manner. Besides, IκBα in cytoplasmic fraction was increased, while p65 level in nuclear fraction was decreased after SAMC treatment in OA. CONCLUSIONS: This study showed that SAMC may play a protective role in IL-1ß induced osteoarthritis (OA) model. This effect may be through inhibiting the NF-κB signaling pathway, therefore altering the MMPs/TIMP-1 ratio change which induced type II collagen destruction and decreasing inflammatory cytokine secretion such as TNF-α.

Influence of an n-3 long-chain polyunsaturated fatty acid-enriched diet on experimentally induced synovitis in horses.

Dietary n-3 long-chain polyunsaturated fatty acid (LCPUFA) supplementation has previously been shown to modify joint-related inflammation in several species, although information in the horse is lacking. We investigated whether dietary supplementation with n-3 LCPUFA would modify experimentally induced synovitis in horses. Twelve, skeletally mature, non-pregnant mares were randomly assigned to either a control diet (CONT) or an n-3 long-chain fatty acid-enriched treatment diet (N3FA) containing 40 g/day of n-3 LCPUFA for 91 days. Blood samples taken on days 0, 30, 60 and 90, and synovial fluid collected on days 0 and 90 were processed for lipid composition. On day 91, joint inflammation was stimulated using an intra-articular (IA) injection of 100 ng of recombinant equine IL-1beta (reIL-1ß). Synovial fluid samples taken at post-injection hours (PIH) 0, 4, 8 and 24 were analysed for prostaglandin E2 (PGE2 ), matrix metalloproteinase (MMP) activity and routine cytology. Synovium and articular cartilage samples collected at PIH 8 were analysed for gene expression of MMP 1 and MMP 13, interleukin-1beta (IL-1ß), cyclooxygenase 2 (COX-2), tumour necrosis factor-alpha and the aggrecanases, a disintegrin and metalloprotease with thrombospondin motifs (ADAMTS)-4 and ADAMTS-5. A 90-day feeding period of n-3 LCPUFA increased serum phospholipid and synovial fluid lipid compositions of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) compared to CONT horses. The reIL-1ß injection caused an inflammatory response; however, there was no effect of dietary treatment on synovial fluid PGE2 content and MMP activity. Synovial tissue collected from N3FA horses exhibited lower expression of ADAMTS-4 compared to CONT horses. Despite the presence of EPA and DHA in the synovial fluid of N3FA horses, dietary n-3 LCPUFA supplementation did not modify synovial fluid biomarkers compared to CONT horses; however, the lower ADAMTS-4 mRNA expression in N3FA synovium warrants further investigation of n-3 LCPUFA as a joint therapy.

Multitargeted effects of hangeshashinto for treatment of chemotherapy-induced oral mucositis on inducible prostaglandin E2 production in human oral keratinocytes.

OBJECTIVE: Chemotherapy-induced oral mucositis (COM) is characterized by painful inflammation with prolonged damage that involves the pathological pain-evoking prostaglandin E2 (PGE2). We previously found that gargling with hangeshashinto (HST), a traditional Japanese medicine, was effective for the treatment of COM. However, little is known regarding the mechanisms. Our aim was to identify the active ingredients and clarify the characteristic effects of HST on the PGE2 system. METHODS: Prostanoids produced by human oral keratinocytes (HOK) stimulated with IL-1ß were measured by enzyme immunoassay. Active ingredients that regulate PGE2 production were identified and quantified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and a culture system of HOK cells. RESULTS: Inducible PGE2, PGD2, and PGF2α, metabolites of cyclooxygenase (COX) pathways, were reduced by HST (10-300 µg/mL) without inducing cytotoxicity. The active ingredients of HST were quantified by LC-MS/MS, and [6]-shogaol, [6]-gingerol, wogonin, baicalein, baicalin, and berberine were shown to reduce PGE2 production. A mixture of these 6 ingredients at concentrations equal to 300 µg/mL of HST strongly suppressed PGE2 production to the same level as HST. [6]-Shogaol and [6]-gingerol did not decrease COX-2 mRNA expression and mostly inhibited PGE2 metabolic activity in an assay using intact HOK cells, suggesting that they regulate PGE2 synthesis at the posttranscriptional level. Wogonin, baicalin, and berberine inhibited expression of COX-2 mRNA without affecting PGE2 metabolic activity. Moreover, wogonin, but not [6]-shogaol, suppressed phosphorylation of mitogen-activated protein kinases (p38s and JNKs). CONCLUSIONS: These lines show that HST includes several PGE2-regulating ingredients that have different mechanisms and can function as a multicomponent and multitarget agent for treatment of COM, indicating that HST may be beneficial in a new medical strategy for COM treatment.

Picralima nitida seeds suppress PGE2 production by interfering with multiple signalling pathways in IL-1ß-stimulated SK-N-SH neuronal cells.

ETHNOPHARMACOLOGICAL RELEVANCE: The dried seed of Picralima nitida is used in rheumatic fever and as an antipyretic in West Africa. In this study we have investigated the effects of an extract obtained from the seeds of Picralima nitida (PNE) on PGE2 production in IL-1ß-stimulated cells. MATERIALS AND METHODS: Prostaglandin E2 (PGE2) was measured in supernatants of IL-1ß-stimulated SK-N-SH cells using enzyme immunoassay (EIA) for PGE2. In Cell ELISA and western blot were used to evaluate the effects of PNE on protein expressions of COX-2, mPGES-1, IκB and IKK. To determine the effect of the extract on NF-κB transactivation, a reporter gene assay was carried out in HEK293 cells stimulated with TNFα. An ELISA was used to measure the roles of p38, ERK1/2 and JNK Mitogen Activated Protein Kinases (MAPKs) on anti-neuroinflammatory actions of PNE. RESULTS: Results show that PNE significantly inhibited PGE2 production, as well as COX-2 and mPGES-1 protein expressions in IL-1ß-stimulated SK-N-SH cells. Molecular targeting experiments showed that PNE interfered with NF-κB signalling pathway through attenuation of TNFα-stimulated NF-κB transcriptional activation in HEK 293 cells. Furthermore, IL-1ß-mediated phosphorylation of IκB and IKK were inhibited in SK-N-SH cells. PNE (50-200 µg/ml) also produced significant inhibition of IL-1ß-induced p38 MAPK phosphorylation in SK-N-SH cells. However, phosphorylation of ERK1/2 and JNK MAPKs were achieved at 100 and 200 µg/ml of the extract. CONCLUSIONS: Taken together, these results clearly demonstrate that Picralima nitida suppresses PGE2 production by targeting multiple pathways involving NF-κB and MAPK signalling in IL-1ß-stimulated SK-N-SH neuronal cells.

L-arginine abolishes the hypothalamic serotonergic activation induced by central interleukin-1ß administration to normal rats.

IL-1ß-induced anorexia may depend on interactions of the cytokine with neuropeptides and neurotransmitters of the central nervous system control of energy balance and serotonin is likely to be one catabolic mediator targeted by IL-1ß. In the complex interplay involved in feeding modulation, nitric oxide has been ascribed a stimulatory action, which could be of significance in counteracting IL-1ß effects.The present study aims to explore the participation of the nitric oxide and the serotonin systems on the central mechanisms induced by IL-1ß and the relevance of their putative interactions to IL-1ß hypophagia in normal rats.Serotonin levels were determined in microdialysates of the ventromedial hypothalamus after a single intracerebroventricular injection of 10 ng of IL-1ß , with or without the pre-injection of 20 µg of the nitric oxide precursor L-arginine. IL-1ß significantly stimulated hypothalamic serotonin extracellular levels, with a peak variation of 130 ± 37% above baseline. IL- 1ß also reduced the 4-h and the 24-h food intakes (by 23% and 58%, respectively). The IL-1ß-induced serotonergic activation was abolished by the pre-injection of L-arginine while the hypophagic effect was unaffected.The data showed that one central effect of IL-1ß is serotonergic stimulation in the ventromedial hypothalamus, an action inhibited by nitric oxide activity. It is suggested that, although serotonin participates in IL-1ß anorexia, other mechanisms recruited by IL-1ß in normal rats are able to override the absence of the serotonergic hypophagic influence.

Fractalkine signaling in microglia contributes to ectopic orofacial pain following trapezius muscle inflammation.

Fractalkine (FKN) signaling is involved in mechanical allodynia in the facial skin following trapezius muscle inflammation. Complete Freund's adjuvant (CFA) injection into the trapezius muscle produced mechanical allodynia in the ipsilateral facial skin that was not associated with facial skin inflammation and resulted in FKN but not FKN receptor (CX3CR1) expression, and microglial activation was enhanced in trigeminal spinal subnucleus caudalis (Vc) and upper cervical spinal cord (C1-C2). Intra-cisterna magna anti-CX3CR1 or anti-interleukin (IL)-1ß neutralizing antibody administration decreased the enhanced excitability of Vc and C1-C2 neurons in CFA-injected rats, whereas intra-cisterna magna FKN administration induced microglial activation and mechanical allodynia in the facial skin. IL-1ß expression and p38 mitogen-activated protein kinase phosphorylation were enhanced in activated microglia after CFA injection. The excitability of neurons whose receptive fields was located in the facial skin was significantly enhanced in CFA-injected rats, and the number of cells expressing phosphorylated extracellular signal-regulated kinase (pERK) following noxious mechanical stimulation of the facial skin was significantly increased in Vc and C1-C2. We also observed mechanical allodynia of the trapezius muscle as well as microglial activation and increased pERK expression in C2-C6 after noxious stimulation of the trapezius muscle in facial skin-inflamed rats. These findings suggest that FKN expression was enhanced in Vc and C1-C2 or C2-C6 following trapezius muscle or facial skin inflammation, microglia are activated via FKN signaling, IL-1ß is released from the activated microglia, and the excitability of neurons in Vc and C1-C2 or C2-C6 is enhanced, resulting in the ectopic mechanical allodynia.

Antioxidant protection of the brain in rats during acute stress and administration of interleukin-1ß.

We studied the effect of IL-1ß on antioxidant enzyme activity in emotiogenic structures of the brain (hypothalamus, sensorimotor cortex, and amygdala) in behaviorally passive and active rats with different sensitivity to stress. One-hour immobilization of animals with simultaneous electrocutaneous stimulation was used as a model of stress. An intraperitoneal injection of IL-1ß (5 µg/kg) was followed by the decrease in glutathione reductase activity in the hypothalamus of rats. Behaviorally active animals of the IL-1ß group were characterized by an increase in the activities of Cu/Zn superoxide dismutase and glutathione peroxidase in the sensorimotor cortex and amygdala, respectively. IL-1ß administration was accompanied by activation of Cu/Zn superoxide dismutase and glutathione peroxidase in the amygdala of passive rats. Pretreatment with IL-1ß abolished the poststress changes in enzyme activity in the hypothalamus and sensorimotor cortex of active and passive rats, respectively. These data illustrate the specific effects of IL-1ß on antioxidant protection of CNS tissues in rats with various behavioral characteristics.

Antiplatelet effect of phloroglucinol is related to inhibition of cyclooxygenase, reactive oxygen species, ERK/p38 signaling and thromboxane A2 production.

Platelet dysfunction is a major risk factor of cardiovascular diseases such as atherosclerosis, stroke and myocardial infarction. Many antiplatelet agents are used for prevention and treatment of these diseases. In this study, phloroglucinol (2.5-25 µM) suppressed AA-induced platelet aggregation and thromboxane B(2) (TXB(2)) production, but not U46619-induced platelet aggregation. Phloroglucinol (100-250 µM) showed little cytotoxicity to platelets. Phloroglucinol inhibited the COX-1 and COX-2 activities by 45-74% and 49-72% respectively at concentrations of 10-50 µM. At concentrations of 1 and 5 µM, phloroglucinol attenuated the AA-induced ROS production in platelets by 30% and 53%, with an IC(50) of 13.8 µM. Phloroglucinol also inhibited the PMA-stimulated ROS production in PMN. Preincubation of platelets by phloroglucinol (10-25 µM) markedly attenuated the AA-induced ERK and p38 phosphorylation. Intravenous administration of phloroglucinol (2.5 and 5 µmol/mouse) suppressed the ex vivo AA-induced platelet aggregation by 57-71%. Phloroglucinol administration also elevated the mice tail bleeding time. Moreover, phloroglucinol inhibited the IL-1ß-induced PGE(2) production in pulp fibroblasts. These results indicate that antiplatelet and anti-inflammatory effects of phloroglucinol are related to inhibition of COX, ROS and TXA2 production as well as ERK/p38 phosphorylation in platelets. Phloroglucinol further suppress PMA-induced ROS production in PMN. The antiplatelet effect of phloroglucinol was confirmed by ex vivo study. Clinically, the consumption of phloroglucinol-containing food/natural products as nutritional supplement may be helpful to cardiovascular health. Phloroglucinol has potential pharmacological use.

Central injection of exogenous IL-1ß in the control activities of hypothalamic-pituitary-gonadal axis in anestrous ewes.

This study was performed to determine the effect of intracerebroventricular (icv) injection of interleukin (IL)-1ß on the gene expression, translation and release of gonadotropin-releasing hormone (GnRH) and the GnRH receptor (GnRHR) gene expression in the hypothalamus of anestrous ewes. In the anterior pituitary gland (AP), the expression of genes encoding: GnRHR, ß subunits of luteinizing hormone (LH) and folliculotropic hormone (FSH) was determined as well as the effect of IL-1ß on pituitary gonadotropins release. The relative mRNA level was determined by real-time PCR, GnRH concentration in the cerebrospinal fluid (CSF) was assayed by ELISA and the plasma concentration of LH and FSH were determined by radioimmunoassay. Our results showed that icv injection of IL-1ß (10 or 50 µg/animal) decreased the GnRH mRNA level in the pre-optic area (POA) (35% and 40% respectively; p ≤ 0.01) and median eminence (ME) (75% and 70% respectively; p ≤ 0.01) and GnRHR gene expression in ME (55% and 50% respectively; p ≤ 0.01). A significant decrease in GnRHR mRNA level in the AP in the group treated with the 50 µg (60%; p ≤ 0.01) but not with the 10 µg dose was observed. The centrally administrated IL-1ß lowered also GnRH concentration in the CSF (60%; p ≤ 0.01) and reduced the intensity of GnRH translation in the POA (p ≤ 0.01). It was not found any effect of icv IL-1ß injection upon the release of LH and FSH. However, the central injection of IL-1ß strongly decreased the LHß mRNA level (41% and 50%; p ≤ 0.01; respectively) and FSHß mRNA in the case of the 50 µg dose (49%; p ≤ 0.01) in the pituitary of anestrous ewes. These results demonstrate that the central IL-1ß is an important modulator of the GnRH biosynthesis and release during immune/inflammatory challenge.

Polyoxypregnane glycoside from Dregea volubilis extract inhibits IL-1ß-induced expression of matrix metalloproteinase via activation of NF-κB in human chondrocytes.

Interleukin-1ß (IL-1ß) induces the expression of matrix metalloproteinases (MMPs) implicated in cartilage and joint degradation in osteoarthritis (OA) and rheumatoid arthritis (RA). Polyoxypregnane glycoside (PPG), active compound was identified from Dregea volubilis extract by chemical analysis, shown to exert chondroprotective effects in cartilage explant models. However, no studies have been undertaken for the molecular investigation of whether PPG constituents protect the human articular chondrocyte (HAC). In the present studies, HAC was co-treated with IL-1ß and PPG. The expression of MMPs, type II collagen, phosphorylation of mitogen-activated protein kinases (MAPKs) and NF-κB signaling pathway were determined by Western immunoblotting. PPG (6.25-25 µM) decreased the IL-1ß-induced HA release from chondrocyte to culture medium. The mode of action of PPG was likely mediated through inhibiting expression of MMP-1, -3 and -13 in the medium, which was associated with the inhibition of mRNA expression. PPG had no effect on IL-1ß-induced phosphorylation of MAPK pathway. Conversely, PPG decreased phosphorylation of IκB kinase and IκBα degradation. Taken together, these results indicate that PPG may inhibit cartilage degradation in OA and may also be used as nutritional supplement for maintaining joint integrity and function.

Noradrenaline release in rodent tissues is inhibited by interleukin-1ß but is not affected by urotensin II, MCH, NPW and NPFF.

We studied whether noradrenaline release is affected by interleukin-1ß and the neuropeptides urotensin II, melanin-concentrating hormone (MCH), neuropeptide W (NPW) and neuropeptide FF (NPFF). Rodent tissues preincubated with [3H]noradrenaline were superfused, and the effect of peptides on the electrically-evoked tritium overflow ("noradrenaline release") was studied. In mouse brain cortex, interleukin-1ß at 0.3 nM and the prostaglandin E2 analogue sulprostone at 3 nM inhibited noradrenaline release by about 40% the effect of interleukin-1ß developed gradually, whereas the effect of sulprostone occurred promptly. Urotensin II at 0.001-1 µM did not affect noradrenaline release in rat kidney cortex, whereas 0.01 µM angiotensin II increased it (positive control). MCH at 0.01-1 µM did not alter noradrenaline release in the rat brain cortex, and NPW 1 µM did not affect noradrenaline release in the mouse hypothalamus or hippocampus. In each model, 0.1 µM sulprostone inhibited noradrenaline release (positive control). NPFF and the NPFF2 receptor agonist dNPA (1 µM) did not affect noradrenaline release in the mouse atria; the inhibitory effect of the δ opioid receptor agonist 1 µM DPDPE on noradrenaline release in this tissue was not altered by NPFF or dNPA at 0.32 µM but was counteracted by the δ opioid antagonist naltrindole at 0.001 µM. In conclusion, interleukin-1ß inhibits noradrenaline release in the mouse cortex; the effect develops gradually, suggesting that it affects protein biosynthesis. Noradrenergic neurons in various tissues from rodents are devoid of presynaptic receptors for urotensin II, MCH, NPW and NPFF. Finally, an interaction between a δ opioid agonist and NPFF could not be detected.

Biphasic effects of baicalin, an active constituent of Scutellaria baicalensis Georgi, in the spontaneous sleep-wake regulation.

AIM OF THE STUDY: Baicalin is an active compound originating from the root of Scutellaria baicalensis Georgi, which has been used for anti-inflammation, anti-bacteria, anti-hypertension, anti-allergy and sedation since ancient China, though the neuronal mechanisms involved in the sedative effect is still unclear. Baicalin possesses the ability to decrease the expression of pro-inflammatory cytokines and nuclear factor (NF)-κB activity. Furthermore, baicalin has demonstrated an anxiolytic-like effect via activation of γ-aminobutyric acid-A (GABA(A)) receptors. Pro-inflammatory cytokines (e.g. interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF)-α) and the GABAergic system promote sleep. This study was designed to determine whether the GABA(A) receptor activation and/or the suppression of pro-inflammatory cytokines mediate(s) baicalin-induced sleep alterations. MATERIALS AND METHODS: Baicalin was intracerebroventricularly (ICV) administered 20 min either prior to the beginning of the light period or before the onset of the dark period. Electroencephalogram (EEG) and gross body movement were acquired for sleep analysis. Pharmacological blockade of IL-1 and GABA(A) receptors were employed to elucidate the involvements of IL-1 and GABA(A) receptors in baicalin-induced sleep alterations. IL-1ß concentrations obtained after baicalin administration in several distinct brain regions were determined by ELISA. RESULTS: ICV administration of baicalin decreased slow wave sleep (SWS) during the first 2h of the light period. Rapid eye movement sleep (REMS) was not altered. The blockade of IL-1ß-induced SWS enhancement by baicalin suggests that the antagonism of IL-1 receptors is involved in baicalin-induced SWS decrement during the light period. However, IL-1ß concentrations during the light period were not altered after baicalin administration. In contrast, baicalin increased both SWS and REMS during hours 8-10 of the dark (active) period when baicalin was administered at the beginning of the dark period, and its effects were blocked by the GABA(A) receptor antagonist bicuculline. CONCLUSION: Baicalin exhibits biphasic effects on sleep-wake regulation; the decrease of SWS during the light period and increases of SWS and REMS during the dark period. Inhibition of IL-1 action and enhancement of GABA(A) receptor activity may mediate baicalin's effects during the light and dark period, respectively.

Interleukin-1 receptor antagonist inhibits the release of glutamate, hydroxyl radicals, and prostaglandin E(2) in the hypothalamus during pyrogen-induced fever in rabbits.

The present study was attempted to determine whether interleukin-1 receptor antagonist (IL-1ra) pretreatment exerts its antipyresis by reducing organum vasculosum laminae terminalis (OVLT) release of glutamate, hydroxyl radicals and prostaglandin E(2) in rabbits. It was found that systemic administration of lipopolysaccharide induced increased levels of both core temperature and OVLT levels of glutamate, hydroxyl radicals, and prostaglandin E(2). The rise in both the core temperature and OVLT glutamate, hydroxyl radicals and prostaglandin E(2) could also be induced by intracerebroventricular injection of interleukin-1beta. Pretreatment with an intracerebroventricular dose of IL-1ra significantly prevented the lipopolysaccharide or IL-1beta-induced overproduction of glutamate, hydroxyl radicals, and prostaglandin E(2) in OVLT of rabbit's brain. The febrile response caused by systemic administration of lipopolysaccharide or central injection of interleukin-1beta could also be IL-1ra pretreatment-ameliorated. These results indicate that IL-1ra pretreatment may exert its antipyresis by inhibiting the glutamate-hydroxyl radicals-prostaglandin E(2) pathways in the OVLT of rabbit's brain during lipopolysaccharide fever.

Evaluation of inflammatory responses induced via intra-articular injection of interleukin-1 in horses receiving a dietary nutraceutical and assessment of the clinical effects of long-term nutraceutical administration.

OBJECTIVE: To evaluate inflammatory responses induced via intra-articular recombinant human interleukin (IL)-1beta treatment in horses receiving a dietary nutraceutical (DN; composed of mussel, shark cartilage, abalone, and Biota orientalis lipid extract) and assess the clinical effects of long-term DN administration. ANIMALS: 22 healthy horses. PROCEDURES: 12 horses were fed 0, 15, 45, or 75 mg of DN (3 horses/treatment) daily for 84 days. General health and clinicopathologic variables were monitored at intervals. Ten other horses received 0 or 15 g of DN/d (5 horses/treatment) for 29 days (beginning day -14). One intercarpal joint in each horse was injected twice with IL-1beta (10 and 100 ng on days 0 and 1, respectively), and the contralateral joint was similarly injected with saline (0.9% NaCl) solution. Synovial fluid prostaglandin E(2) (PGE(2)), sulfated glycosaminoglycan (GAG), nitric oxide (NO), and protein concentrations and leukocyte counts were analyzed before and at intervals after injections. RESULTS: Administration of the DN (up to 75 g/d) to horses for 84 days did not induce any adverse effects. In the other experiment, synovial fluid PGE(2), GAG, and protein concentrations and leukocyte count increased after intra-articular injections of IL-1beta (compared with effects of saline solution injections) in horses that received no DN; NO concentration was not affected. In horses that were fed the DN, intra-articular IL-1beta injections did not induce significant increases in synovial fluid PGE(2) and GAG concentrations. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that administration of the DN may be useful in preventing inflammation associated with arthritis and degenerative joint disease in horses.

Interleukin-1beta and insulin elicit different neuroendocrine responses to hypoglycemia.

Interleukin (IL)-1beta induces a prolonged hypoglycemia in mice that is not caused by a reduction in food intake and is dissociable from insulin effects. There is a peripheral component in the hypoglycemia that the cytokine induces resulting from an increased glucose uptake, an effect that can be exerted in a paracrine fashion at the site where IL-1 is locally produced. However, the maintenance of hypoglycemia is controlled at brain levels because the blockade of IL-1 receptors in the central nervous system inhibits this effect to a large extent. Furthermore, there is evidence that the cytokine interferes with counter regulation to hypoglycemia. Here we report that administration of IL-1 or long-lasting insulin results in different changes in food intake and in neuroendocrine mechanisms 8 h following induction of the same degree of hypoglycemia (40-45% decrease in glucose blood levels). Insulin, but not IL-1, caused an increase in food intake and an endocrine response that tends to reestablish euglycemia. Conversely, a decrease in noradrenergic and an increase in serotonergic activity in the hypothalamus occur in parallel with a reduction of glucose blood levels only in IL-1-treated mice, effects that can contribute to the maintenance of hypoglycemia. These results are compatible with the proposal that IL-1 acting in the brain can reset glucose homeostasis at a lower level. The biologic significance of this effect is discussed.

Administration of IL-1beta to the 4th ventricle causes anorexia that is blocked by agouti-related peptide and that coincides with activation of tyrosine-hydroxylase neurons in the nucleus of the solitary tract.

Inflammation-associated cachexia is associated with multiple chronic diseases and involves activation of appetite regulating centers in the arcuate nucleus of the hypothalamus (ARH). The nucleus of the solitary tract (NTS) in the brainstem has also been implicated as an important nucleus involved in appetite regulation. We set out to determine whether the NTS may be involved in inflammation-associated anorexia by injecting IL-1 beta into the 4th ventricle and assessing food intake and NTS neuronal activation. Injection of IL-1 beta produced a decrease in food intake at 3 and 12h after injection which was ameliorated at the 12h time point by a sub-threshold dose of agouti-related peptide (AgRP). Investigation into neuron types in the NTS revealed that IL-1 beta injection was associated with an increase in c-Fos activity in NTS neurons expressing tyrosine hydroxylase (TH). Additionally, injection of IL-1 beta into the 4th ventricle did not produce c-Fos activation of neurons expressing pro-opiomelanocortin (POMC) in the ARH, cells known to be involved in producing anorexia in response to systemic inflammation. Double-label in situ hybridization revealed that TH neurons did not express IL-1 receptor I (IL1-RI) transcript, demonstrating that c-Fos activation of TH neurons in this setting was not via direct stimulation of IL-1 beta on TH neurons themselves. We conclude that IL-1 beta injection into the 4th ventricle produces anorexia and is accompanied by an increase in activation in TH neurons in the NTS. This provides evidence that the brainstem may be an important mediator of anorexia in the setting of inflammation.

[Recombinant human interleukine-1beta (betaleukine) usage for acute radiation sickness of severe degree treatment at canines].

Recombinant human interleukine-1beta (betaleukune) of Institute of especially pure biopreparation production had been examined as a treatment means at acute radiation disease of severe degree at dogs. Dogs were irradiated totally in doses above the LD95/45. Betaleukine had been administered s/c twice in day in 15 min - 2 h after irradiation. All the dogs, including control ones, received in acute period 8-24/26 days after irradiation antibiotics ampicillin and gentamycin i/m. Betaleukine increased 45 day-survival by 37% at 4 Gy and by 25% at 4.4 Gy. The effect correlated with more high level of nadir and more early beginning the leucocyte number restoration. We observed the regularity at all the dogs that received betaleukine as survived as died, but in the latter case in a lesser degree certainly. Besides the noticed character of leucocytes kinetics had been repeated at all the blood cell types but in the different degree. It had been concluded on the base of these observations that betaleukine acts on hemopoietic stem cells preferentially. The effect is in preventing death of a stem cells part, or in stimulating survived stem cells proliferation, or in both together. Betaleukin can be regarded as a suitable means of urgent pathogenic therapy at radiation accidents.