Antioxidant, anti-inflammatory and neuroprotective actions of resveratrol after experimental nervous system insults. Special focus on the molecular mechanisms involved.

After different types of acute central nervous system insults, including stroke, subarachnoid haemorrhage and traumatic brain and spinal cord injuries, secondary damage plays a central role in the induction of cell death, neurodegeneration and functional deficits. Interestingly, secondary cell death presents an attractive target for clinical intervention because the temporal lag between injury and cell loss provides a potential window for effective treatment. While primary injuries are the direct result of the precipitating insult, secondary damage involves the activation of pathological cascades through which endogenous factors can exacerbate initial tissue damage. Secondary processes, usually interactive and overlapping, include oxidative stress, neuroinflammation and dysregulation of autophagy, ultimately leading to cell death. Resveratrol, a natural stilbene present at relatively high concentrations in grape skin and red wine, exerts a wide range of beneficial health effects. Within the central nervous system, in addition to its inherent free radical scavenging role, resveratrol increases endogenous cellular antioxidant defences thus modulating multiple synergistic pathways responsible for its antioxidant, anti-inflammatory and anti-apoptotic properties. During the last years, a growing body of in vitro and in vivo evidence has been built, indicating that resveratrol can induce a neuroprotective state and attenuate functional deficits when administered acutely after an experimental injury to the central nervous system. In this review, we summarize the most recent findings on the molecular pathways involved in the neuroprotective effects of this multi target polyphenol, and discuss its neuroprotective potential after brain or spinal cord injuries.

Intrathecal Administration of an Anti-nociceptive Non-CpG Oligodeoxynucleotide Reduces Glial Activation and Central Sensitization.

Inflammatory pain associates with spinal glial activation and central sensitization. Systemic administration of IMT504, a non-CpG oligodeoxynucleotide originally designed as an immunomodulator, exerts remarkable anti-allodynic effects in rats with complete Freund´s adjuvant (CFA)-induced hindpaw inflammation. However, the anti-nociceptive mechanisms of IMT504 remain unknown. Here we evaluated whether IMT504 blocks inflammatory pain-like behavior by modulation of spinal glia and central sensitization. The study was performed in Sprague Dawley rats with intraplantar CFA, and a single lumbosacral intrathecal (i.t.) administration of IMT504 or vehicle was chosen to address if changes in glial activation and spinal sensitization relate to the pain-like behavior reducing effects of the ODN. Naïve rats were also included. Von Frey and Randall-Selitto tests, respectively, exposed significant reductions in allodynia and mechanical hypersensitivity, lasting at least 24 h after i.t. IMT504. Analysis of electromyographic responses to electrical stimulation of C fibers showed progressive reductions in wind-up responses. Accordingly, IMT504 significantly downregulated spinal glial activation, as shown by reductions in the protein expression of glial fibrillary acidic protein, CD11b/c, Toll-like receptor 4 (TLR4) and the phosphorylated p65 subunit of NFκB, evaluated by immunohistochemistry and western blot. In vitro experiments using early post-natal cortical glial cultures provided further support to in vivo data and demonstrated IMT504 internalization into microglia and astrocytes. Altogether, our study provides new evidence on the central mechanisms of anti-nociception by IMT504 upon intrathecal application, and further supports its value as a novel anti-inflammatory ODN with actions upon glial cells and the TLR4/NFκB pathway. Intrathecal administration of the non-CpG ODN IMT504 fully blocks CFA-induced mechanical allodynia and hypersensitivity, in association with reduced spinal sensitization. Administration of the ODN also results in downregulated gliosis and reduced TLR4-NF-κB pathway activation. IMT504 uptake into astrocytes and microglia support the concept of direct modulation of CFA-induced glial activation.

Resveratrol exerts anti-oxidant and anti-inflammatory actions and prevents oxaliplatin-induced mechanical and thermal allodynia.

Chemotherapy-induced peripheral neuropathic pain (CIPNP) is a frequent and devastating side effect of cancer therapy. No preventive strategies are currently available. We investigated the use of resveratrol (RESV) in the prevention of CIPNP and evaluated key components of the antioxidant defense system and neuroinflammatory factors as possible mediators contributing to RESV actions. Male rats were injected with oxaliplatin (OXA) and received daily oral RESV. Paw mechanical and thermal allodynia, oxidative stress, antioxidant, pro-inflammatory and neuronal injury/activation markers were evaluated in the sciatic nerve (SN), lumbar dorsal root ganglia (DRG) and spinal cord (SC). OXA-injected animals developed mechanical and thermal allodynia, while those receiving OXA + RESV showed patterns of response similar to control animals. Higher TBARS levels and lower GSH/GSSG ratios were observed in the SN of animals receiving OXA. The mRNA levels of the transcription factor NFκB and the pro-inflammatory cytokine TNFα were found to be upregulated both in lumbar DRG and SC. In addition, the antioxidant enzymes NQO-1 and HO-1 and the neuronal injury marker ATF3 showed increased levels of expression in lumbar DRG. In the dorsal SC the neuronal activation marker c-fos and the transcription factor Nrf2, main regulator of antioxidant defenses, were found to be upregulated. RESV early and sustained administration prevented NFκB, TNFα, ATF3 and c-fos upregulation, while increasing the expression of Nrf2, NQO-1, HO-1 and the redox-sensitive deacetylase SIRT1. RESV treatment was also able to restore TBARS levels and GSH/GSSG ratio. Thus, RESV administration resulted in the upregulation of antioxidant mediators, suppression of pro-inflammatory parameters and prevention of OXA-induced mechanical and thermal allodynia.

Progesterone reduces the expression of spinal cyclooxygenase-2 and inducible nitric oxide synthase and prevents allodynia in a rat model of central neuropathic pain.

BACKGROUND: Spinal cord injury (SCI) results in the development of chronic pain that is refractory to conventional treatment. Progesterone, a neuroprotective steroid, may offer a promising perspective in pain modulation after central injury. Here, we explore the impact of progesterone administration on the post-injury inflammatory cascade involving the enzymes cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) at the spinal cord level. We also analyse pain behaviours, the profile of glial cell activation, and IκB-α mRNA levels, as an index of NF-κB transactivation. METHODS: We used biochemical, immunohistochemical and molecular techniques, as well as behavioural studies, to investigate the effects of progesterone in a well-characterized model of central neuropathic pain. RESULTS: Injured animals receiving progesterone presented reduced mRNA levels of the proinflammatory enzymes, as well as decreased COX-2 activity and nitrite levels, as compared to vehicle-treated injured rats. Further, animals receiving the steroid exhibited lower levels of IκB-α mRNA, suggesting decreased NF-κB transactivation. Progesterone administration also attenuated the injury-induced increase in the number of glial fibrillary acidic protein and OX-42 positive cells both at early and late time points after injury, and prevented the development of mechanical and thermal allodynia. Further, when injured rats received early progesterone administration for a critical period of time after injury, they did not display allodynic behaviours even after the treatment had stopped. CONCLUSIONS: Our results suggest that progesterone, by modulating early neuroinflammatory events triggered after SCI, may represent a useful strategy to prevent the development of central chronic pain.

Bone marrow stromal cells attenuate injury-induced changes in galanin, NPY and NPY Y1-receptor expression after a sciatic nerve constriction.

Single ligature nerve constriction (SLNC) of the rat sciatic nerve triggers neuropathic pain-related behaviors and induces changes in neuropeptide expression in primary afferent neurons. Bone marrow stromal cells (MSCs) injected into the lumbar 4 (L4) dorsal root ganglia (DRGs) of animals subjected to a sciatic nerve SLNC selectively migrate to the other ipsilateral lumbar DRGs (L3, L5 and L6) and prevent mechanical and thermal allodynia. In this study, we have evaluated the effect of MSC administration on the expression of the neuropeptides galanin and NPY, as well as the NPY Y(1)-receptor (Y(1)R) in DRG neurons. Animals were subjected to a sciatic nerve SLNC either alone or followed by the administration of MSCs, phosphate-buffered saline (PBS) or bone marrow non-adherent mononuclear cells (BNMCs), directly into the ipsilateral L4 DRG. Seven days after injury, the ipsilateral and contralateral L4-5 DRGs were dissected out and processed for standard immunohistochemistry, using specific antibodies. As previously reported, SLNC induced an ipsilateral increase in the number of galanin and NPY immunoreactive neurons and a decrease in Y(1)R-positive DRG neurons. The intraganglionic injection of PBS or BNMCs did not modify this pattern of expression. In contrast, MSC administration partially prevented the injury-induced changes in galanin, NPY and Y(1)R expression. The large number of Y(1)R-immunoreactive neurons together with high levels of NPY expression in animals injected with MSCs could explain, at least in part, the analgesic effects exerted by these cells. Our results support MSC participation in the modulation of neuropathic pain and give insight into one of the possible mechanisms involved.

Nitric oxide, superoxide, and hydrogen peroxide production in brain mitochondria after haloperidol treatment.

Inhibition of mitochondrial respiration and free radical induction have been suggested to be involved in haloperidol neurotoxicity. In this study, mice were injected i.p. with haloperidol, according to two different treatments: (a) a single injection (1 mg/kg), sacrificed 1 h after the injection (single-dose model); and (b) two injections (1 mg/kg each), sacrificed 24 h after the first dose (double-dose model). Determinations of oxygen consumption and hydrogen peroxide (H2O2) production rate were carried out in isolated brain mitochondria. Nitric oxide (NO) and superoxide (O2-) production rates were measured in submitochondrial particles (SMP). Single-dose haloperidol treatment produced a 33% inhibition in malate-glutamate-dependent respiration, while no significant changes were found after double-dose treatment. NO production was inhibited by 39 and 54% in SMP from haloperidol-treated mice (single- and double-dose treatments, respectively) (control value: 1.6 +/- 0.2 nmol/min mg protein). NO steady-state concentration was estimated at about 16.5 nM and was decreased by 40% by haloperidol treatment. Increases of 105 and 54% were found in succinate-supported O2- and H2O2 production rates, respectively, after haloperidol single-dose treatment. Haloperidol treatment generated a 248% increase in SMP O2- production rate when measured in the presence of NADH plus rotenone. Our results suggest that haloperidol neurotoxicity would be mediated by a decreased mitochondrial NO production, a decreased intramitochondrial NO steady-state concentration, and by an inhibition of mitochondrial electron transfer with enhancement of O2- and H2O2 production. This inhibition does not seem to be caused by increased NO or ONOO- formation.