Evaluation of the Gastroprotective Effect of the Flavonoid Complex from Lychnis chalcedonica L. on the Models of Experimental Ulcerogenesis.

Using rat and mouse models of neurogenic, ethanol-induced, and indometacin-induced damage to the gastric mucosa we demonstrated that course preventive treatment with flavonoid complex from aerial parts of Lychnis chalcedonica L. increased the resistance of gastric mucosa to ulcerogenic factors of different etiology. The gastroprotective effect of the phytocomplex in a dose range of 16-1600 µg/kg was comparable with that of the reference drug plantaglucide and was superior to that of the reference drugs eleutherococcus extract and methyluracil in the therapeutic doses. The antiulcerogenic activity of Lychnis chalcedonica flavonoid complex considerably exceeded activity of Lychnis chalcedonica L. extract demonstrated in our previous experiments.

Fish oil supplementation attenuates neuroinflammation and alleviates depressive-like behavior in rats submitted to repeated lipopolysaccharide.

PURPOSE: Depression is frequently associated with inflammation, whereas omega-3 polyunsaturated fatty acids (PUFAs) primarily found in fish oil possess anti-inflammatory properties. Although converging studies suggest an antidepressant effect of PUFAs, there is limited evidence directly linking the neuro-immune modulating features of PUFAs to the antidepressant actions. METHODS: Therefore, we assessed the effects of fish oil (FO) supplementation on behavioral changes, inflammatory cytokine expression and oxidative reactions in frontal cortex and hippocampus of rats following repeated peripheral immune challenge by lipopolysaccharide (LPS) for 2 weeks (500 µg/kg every other day). RESULTS: Repeated LPS administration induced the rats to a depressive-like state and increased mRNA expression of pro-inflammatory cytokines, including 1L-1ß, 1L-6 and TNF-α, in frontal cortex and hippocampus. FO supplementation attenuated the LPS-induced abnormal behavior and brain inflammatory response. Concurrent with the antidepressant action, FO also reduced LPS-induced oxidative reactions and neural apoptosis in the rat brain, as evidenced by decreased malondialdehyde (MDA) production, increased catalase activities and inhibited pro-apoptotic protein Bax mRNA expression. In addition, FO inhibited activation of NF-κB and iNOS induced by LPS. Interestingly, we found FO suppressed the activation of the inflammasome NLRP3 and ionotropic purinergic receptor P2X7R evoked by LPS, suggesting a potential anti-inflammatory mechanism for PUFAs. Besides, FO also restored the LPS-induced neurochemical disturbance, especially the balance between serotonin and kynurenine branches of tryptophan metabolism, which is tightly associated with depression. CONCLUSIONS: These findings provide novel insights into the antidepressant action of PUFAs and further strengthen the link between inflammation and depression.

Wogonin prevents TLR4-NF-κB-medicated neuro-inflammation and improves retinal ganglion cells survival in retina after optic nerve crush.

Chronic neuro-inflammation is involved in the death of retinal ganglion cells (RGCs) in glaucoma. The aim of this study is to determine whether wogonin can suppress inflammatory responses and rescue RGCs death after optic nerve crush (ONC), an ideal animal model of glaucoma. Wogonin was administered intraperitoneally 10 min after establishment of ONC model. In this study, wogonin treatment reduced RGCs loss and inhibited RGCs apoptosis demonstrated by the increased Brn3a labeling RGCs at day 14 and the decreased cleaved caspase-3 expression at day 7 after ONC, respectively. In ONC model, number of GFAP-positive glial cells and iba1-positive microglial cells were increased, combined of the elevated level of pro-inflammatory cytokines released in retina at day 7. However, most of these responses were inhibited after wogonin treatment. The level of TLR4 expression, NF-κB-P65 nucleus location and NF-κB-P65 phosphorylation were increased in retina at day 1 after ONC, which was significantly reduced after wogonin treatment. These results demonstrated that wogonin protected RGCs survival and suppressed neuro-inflammation in retina after ONC by inhibiting TLR4-NF-κB pathways. We conclude that wogonin could be a possible strategy for the treatment of glaucoma.

Role of capsaicin-sensitive nerves and tachykinins in mast cell tryptase-induced inflammation of murine knees.

OBJECTIVE, DESIGN: Mast cell tryptase (MCT) is elevated in arthritic joints, but its direct effects are not known. Here, we investigated MCT-evoked acute inflammatory and nociceptive mechanisms with behavioural, in vivo imaging and immunological techniques. MATERIAL AND SUBJECTS: Neurogenic inflammation involving capsaicin-sensitive afferents, transient receptor potential vanilloid 1 receptor (TRPV1), substance P (SP), neurokinin A (NKA) and their NK1 tachykinin receptor were studied using gene-deleted mice compared to C57Bl/6 wildtypes (n = 5-8/group). TREATMENT: MCT was administered intraarticularly or topically (20 µl, 12 µg/ml). Capsaicin-sensitive afferents were defunctionalized with the TRPV1 agonist resiniferatoxin (RTX; 30-70-100 µg/kg s.c. pretreatment). METHODS: Knee diameter was measured with a caliper, synovial perfusion with laser Doppler imaging, mechanonociception with aesthesiometry and weight distribution with incapacitance tester over 6 h. Cytokines and neuropeptides were determined with immunoassays. RESULTS: MCT induced synovial vasodilatation, oedema, impaired weight distribution and mechanical hyperalgesia, but cytokine or neuropeptide levels were not altered at the 6-h timepoint. Hyperaemia was reduced in RTX-treated and TRPV1-deleted animals, and oedema was absent in NK1-deficient mice. Hyperalgesia was decreased in SP/NKA- and NK1-deficient mice, weight bearing impairment in RTX-pretreated, TRPV1- and NK1-deficient animals. CONCLUSIONS: MCT evokes synovial hyperaemia, oedema, hyperalgesia and spontaneous pain. Capsaicin-sensitive afferents and TRPV1 receptors are essential for vasodilatation, while tachykinins mediate oedema and pain.

Expression levels of neuroimmune biomarkers in hypothalamus of allergic mice after phthalate exposure.

Previously, we demonstrated that maternal exposure to phthalates enhances atopic dermatitis in male mouse offspring. However, whether phthalate exposure affects neuroimmune biomarkers in allergic mice has not yet been studied. Di-(2-ethylhexyl) phthalate (DEHP) and di-isononyl phthalate (DINP) are environmental chemicals that are commonly used as plasticizers. This study was designed to investigate the expression levels of neuroimmune biomarkers in the hypothalamus of a murine model of allergic asthma after phthalate exposure throughout juvenility until adulthood. Six-week-old C3H/HeJ Jcl male mice were treated with DEHP or DINP (0, 0.02, 0.4 or 8 nmol per body per week) and ovalbumin (OVA; 1 µg per body per 2 weeks) for 7 weeks intratracheally. On the day after the completion of the phthalate and OVA treatment, the hypothalamus from each mouse was collected, and the mRNA expression levels of neuroimmune biomarkers were examined using a real-time RT-PCR analysis. The mRNA expression levels of the proinflammatory cytokines interleukin (IL)-1ß and tumor necrosis factor (TNF)-α, the chemokine CCL3, the transcription factor nuclear factor (NF)-κB, the oxidative stress marker heme-oxygenase (HO)1, a nerve growth factor, and the microglia marker Iba1 were remarkably up-regulated in the hypothalami of mice treated with 8 nmol of DEHP in the presence of the allergen. However, no significant changes were observed, except for reductions in the TNF-α and CCL2 mRNA levels, in mice exposed to DINP combined with the allergen. This study is the first report to show that high-dose DEHP exposure throughout juvenility until adulthood may induce neuroinflammation by modulating neuroimmune biomarkers in the hypothalami of allergic mice.

The neuroendocrine-immune connection regulates chronic inflammatory disease in allergy.

Allergy is an instructive model to study neuroendocrine-immune interaction in chronic inflammation, a key research task taken on by a relatively new scientific field: psychoneuroimmunology (PNI). Itch, as the prime symptom of many chronic inflammatory diseases but especially of allergic inflammation, hints at the prominent role of neurogenic inflammation in the course of the disease. Environmental factors ranging from allergens to perceived stress can trigger the release of neuropeptides from peripheral nerve endings that than activate mast cells and induce an exaggerated alarm response in peripheral organs such as the skin. Beyond this innate immune response, neuroendocrine-immune interaction regulates specific immune balance. Depending on intensity and timing of neuroendocrine activation, especially neuropeptides and neurotrophins either enhance or suppress tissue regeneration and inflammation, the conditions of which will be discussed in detail here. Allergic inflammation thus serves to understand the clinical and therapeutic implications of neuroendocrine-immune interaction in chronic inflammatory disease and its implications for future treatment strategies.

The antinociceptive activity of harmicine on chemical-induced neurogenic and inflammatory pain models in mice.

Harmicine is a ß-carboline alkaloid isolated and identified as a major active compound present in many plant species and marine invertebrates. This alkaloid exhibits a wide spectrum of pharmacological activities, including antispasmodic, antipyretic, and anticancer properties. This report described the antinociceptive properties of harmicine by means of chemical experimental models in order to evaluate the use for pain relief. The results demonstrating the potential analgesic properties of harmicine administered intraperitoneally were shown with the writhing test, reducing writhes around 60% (1 mg/kg), and in the formalin test, where harmicine was more effective toward neurogenic (reducing reaction time around 60%, 1 mg/kg) than inflammatory (68% reduction, 10 mg/kg) pain responses. Furthermore, these effects may operate via vanilloid receptors as revealed by the capsaicin test (41% reduction, with 3 mg/kg), as well as via peripheral glutamate receptors as shown by the glutamate test (50% reduction, with 1 mg/kg). Moreover, the opioid antagonist naloxone hydrochloride did not interfere in the antinociceptive properties of harmicine in the writhing test, revealing that this effect may not have a relationship with the opioid systems. Concluding, this report highlights harmicine as a new candidate to be used as analgesic in the future. Therefore, further studies are being undertaken in order to understand the exact mechanisms involved with the antinociceptive properties of harmicine.

Central nervous system mast cells in peripheral inflammatory nociception.

BACKGROUND: Functional aspects of mast cell-neuronal interactions remain poorly understood. Mast cell activation and degranulation can result in the release of powerful pro-inflammatory mediators such as histamine and cytokines. Cerebral dural mast cells have been proposed to modulate meningeal nociceptor activity and be involved in migraine pathophysiology. Little is known about the functional role of spinal cord dural mast cells. In this study, we examine their potential involvement in nociception and synaptic plasticity in superficial spinal dorsal horn. Changes of lower spinal cord dura mast cells and their contribution to hyperalgesia are examined in animal models of peripheral neurogenic and non-neurogenic inflammation. RESULTS: Spinal application of supernatant from activated cultured mast cells induces significant mechanical hyperalgesia and long-term potentiation (LTP) at spinal synapses of C-fibers. Lumbar, thoracic and thalamic preparations are then examined for mast cell number and degranulation status after intraplantar capsaicin and carrageenan. Intradermal capsaicin induces a significant percent increase of lumbar dural mast cells at 3 hours post-administration. Peripheral carrageenan in female rats significantly increases mast cell density in the lumbar dura, but not in thoracic dura or thalamus. Intrathecal administration of the mast cell stabilizer sodium cromoglycate or the spleen tyrosine kinase (Syk) inhibitor BAY-613606 reduce the increased percent degranulation and degranulated cell density of lumbar dural mast cells after capsaicin and carrageenan respectively, without affecting hyperalgesia. CONCLUSION: The results suggest that lumbar dural mast cells may be sufficient but are not necessary for capsaicin or carrageenan-induced hyperalgesia.

Rostral ventromedial medulla µ, but not κ, opioid receptors are involved in electroacupuncture anti-hyperalgesia in an inflammatory pain rat model.

It has been reported that intracerebroventricular injection of a µ receptor antagonist blocked 2 but not 100Hz electroacupuncture (EA)-produced analgesia in an uninjured animal model. Because persistent pain changes neural response to external stimulation, we hypothesized that the mechanisms of EA anti-hyperalgesia may be different in persistent pain than in health. Hyperalgesia, decreased paw withdrawal latency (PWL) to a noxious thermal stimulus, was induced by subcutaneously injecting complete Freund's adjuvant (CFA) into the hind paws of rats. Selective antagonists against µ (CTOP: D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-ThrNH2, 6.25 nmol) and κ (Nor-BIN: nor-binaltorphimine, 10 nmol) opioid receptors were infused into the rostral ventromedial medulla (RVM) 10 min before a 30-min EA treatment at acupoint Huantiao (GB30) 1h 30 min post-CFA. PWL was measured before and 2.5 post-CFA. Both 10 Hz and 100 Hz EA-produced anti-hyperalgesia were blocked by intra-RVM µ, but not κ, receptor antagonists. Double immunofluorescence staining demonstrated that µ receptor-containing neurons were GABAnergic and that GABAa receptor-containing neurons were serotonergic in the RVM. The results demonstrated an involvement of RVM µ, but not κ, receptors in EA-produced anti-hyperalgesia. In summary, EA may induce release of endogenous endomorphins that activate µ opioid receptors in GABAnergic neurons to suppress the release of GABA. This removes the tonic inhibition of GABA on serotonergic neurons in the RVM, and activation of these serotonergic neurons inhibits pain. EA may be used as complementary treatment for inflammatory pain.

Development of an in vitro coculture of primary sensitive pig neurons and keratinocytes for the study of cutaneous neurogenic inflammation.

Cutaneous neurogenic inflammation (CNI) is often associated with skin disorders. Activated sensory neurons secrete neuropeptides, such as substance P (SP), which initiate or aggravate inflammation in the skin. The discovery of new molecules acting on these neurons is hampered by the difficulty of reproducing the interactions between nerve endings and skin in vitro. We developed an in vitro model based on the coculture of porcine primary keratinocytes and sensory neurons, which mimics skin innervation. To test the relevance of this model, we compared the effects of different substances on CNI by measuring SP secretion in vitro using a sensitive enzyme immunoassay. Collectively, our results indicate that the use of porcine cells could be very useful to perform an in vitro model of CNI. By adding capsaicin, which induces the secretion of SP by neurons, to the culture, we show that our model mimics CNI in vitro, allowing us to screen for molecules that inhibit this inflammatory response. Such a model can be used to test the effects of different substances on CNI and may be useful for dermatological or cosmetic applications. Based on our screen, we found that extracts of Laminaria digitata and Vernonia sublutea inhibit CNI.

Neuroinflammation in Alzheimer's disease: different molecular targets and potential therapeutic agents including curcumin.

Alzheimer's disease (AD) is a neurodegenerative disorder of the elderly. Deposition of amyloid beta plaque and associated neuroinflammation are the major hallmarks of AD. Whereas reactive oxygen species (ROS) and activated microglial cells contribute to neuronal loss, nuclear factor kappaB and apolipoprotein E participate in inflammatory process of AD. Current FDA approved drugs provide only symptomatic relief in AD. For broad spectrum of activity, some natural products are also being tested. Turmeric is used as an anti-inflammatory medicine in various regions of Asia. Curcumin, which is a yellow colored polyphenol compound present in turmeric, showed anti-inflammatory properties. Herein, we discuss the neurobiological and neuroinflammatory pathways of AD, evaluate different molecular targets and potential therapeutic agents, including curcumin, for the treatment of AD.

Effect of pituitary adenylate cyclase activating polypeptide-38 on sensory neuropeptide release and neurogenic inflammation in rats and mice.

Substance P (SP) and calcitonin gene-related peptide (CGRP), released from capsaicin-sensitive sensory nerves induce local neurogenic inflammation, while somatostatin exerts systemic anti-inflammatory actions. The aim of the present study was to investigate the release of pituitary adenylate cyclase activating polypeptide-38 (PACAP-38) and its effects on sensory neuropeptide release in vitro and acute neurogenic ear swelling in vivo. Capsaicin (10(-6) M) or electrical field stimulation (EFS; 40 V, 0.1 ms, 10 Hz, 120 s; 1200 impulses)-induced release of PACAP-38, SP, CGRP and somatostatin from isolated rat tracheae was measured with radioimmunoassay. Mustard oil-induced neurogenic inflammation in the mouse ear was determined with a micrometer and in the rat hind paw skin by the Evans Blue leakage technique. Capsaicin and EFS evoked 27% and more than twofold elevation of PACAP-38 release respectively, compared with the prestimulated basal values from isolated trachea preparation. Exogenously administered PACAP-38 (20-2000 nM) diminished both capsaicin- and EFS-evoked sensory neuropeptide release in a concentration-dependent manner. The maximal inhibitory effects of PACAP on capsaicin-induced substance P, CGRP and somatostatin release amounted to 75.4%, 73.3% and 90.0%, while EFS-evoked release of these peptides was 80.03%, 87.7% and 67.7%. In case of capsaicin stimulation the EC50 values for substance P, CGRP and somatostatin were 82.9 nM, 60.1 nM and 66.9 nM, respectively. When EFS was performed, these corresponding EC50 data were 92.1 nM, 67.8 nM and 20.9 nM. PACAP-38 (10, 100 and 1000 microg/kg i.p. in 200 microl volume) inhibited neurogenic ear swelling in the mouse. Furthermore, 100 microg/kg i.p. PACAP also significantly diminished mustard oil-evoked plasma protein extravasation in the rat skin. These results suggest that PACAP-38 is released from the stimulated peripheral terminals of capsaicin-sensitive afferents and it is able to inhibit the outflow of sensory neuropeptides. Based on this mechanism of action PACAP is also able to effectively diminish/abolish neurogenic inflammatory response in vivo after systemic administration.

Selective ablation of nociceptive neurons for elimination of hyperalgesia and neurogenic inflammation.

OBJECT: Neuropathic pain is mediated by nociceptive neurons that selectively express the vanilloid receptor 1 (VR1). Resiniferatoxin (RTX) is an excitotoxic VR1 agonist that causes destruction of VR1-positive neurons. To determine whether RTX can be used to ablate VR1-positive neurons selectively and to eliminate hyperalgesia and neurogenic inflammation without affecting tactile sensation and motor function, the authors infused it unilaterally into the trigeminal ganglia in Rhesus monkeys. METHODS: Either RTX (three animals) or vehicle (one animal) was directly infused (20 microl) into the right trigeminal ganglion in Rhesus monkeys. Animals were tested postoperatively at 1, 4, and 7 weeks thereafter for touch and pain perception in the trigeminal distribution (application of saline and capsaicin to the cornea). The number of eye blinks, eye wipes, and duration of squinting were recorded. Neurogenic inflammation was tested using capsaicin cream. Animals were killed 4 (one monkey) and 12 (three monkeys) weeks postinfusion. Histological and immunohistochemical analyses were performed. Throughout the duration of the study, response to high-intensity pain stimulation (capsaicin) was selectively and significantly reduced (p < 0.001, RTX-treated compared with vehicle-treated eye [mean +/- standard deviation]): blinks, 25.7 +/- 4.4 compared with 106.6 +/- 20.8; eye wipes, 1.4 +/- 0.8 compared with 19.3 +/- 2.5; and squinting, 1.4 +/- 0.6 seconds compared with 11.4 +/- 1.6 seconds. Normal response to sensation was maintained. Animals showed no neurological deficit or sign of toxicity. Neurogenic inflammation was blocked on the RTX-treated side. Immunohistochemical analysis of the RTX-treated ganglia showed selective elimination of VR1-positive neurons. CONCLUSIONS: Nociceptive neurons can be selectively ablated by intraganglionic RTX infusion, resulting in the elimination of high-intensity pain perception and neurogenic inflammation while maintaining normal sensation and motor function. Analysis of these findings indicated that intraganglionic RTX infusion may provide a new treatment for pain syndromes such as trigeminal neuralgia as well as others.

Repeated subinflammatory ultraviolet B irradiation increases substance P and calcitonin gene-related peptide content and augments mustard oil-induced neurogenic inflammation in the skin of rats.

The cutaneous neurosensory system is suggested to be involved in the pathophysiology of pruritus and skin diseases such as psoriasis. We investigated if repeated subinflammatory doses of ultraviolet B (UVB) irradiation similar to those used to treat pruritus or psoriasis would affect the cutaneous neurosensory system. Sprague-Dawley rats were irradiated thrice weekly for 2-4 weeks with subinflammatory doses of UVB. Three days after the last UVB exposure: (i), the skin contents of substance P (SP), calcitonin gene-related peptide (CGRP), and nerve growth factor (NGF) were quantified; (ii), the skin nerve fiber density was observed; and (iii), the effect of UVB on mustard oil-induced neurogenic inflammation was determined. UV exposure significantly increased SP and CGRP content and mustard oil-induced neurogenic inflammation in UV-irradiated but not non-irradiated skin; however, it did not affect cutaneous NGF content or overall nerve fiber density. These data suggest that repeated subinflammatory UVB irradiation locally increases the content of cutaneous SP and CGRP by an increase of neuropeptide content of nerve fibers rather than by an increase of overall nerve fiber density.