Therapeutic impact of orally administered cannabinoid oil extracts in an experimental autoimmune encephalomyelitis animal model of multiple sclerosis.

There is a growing surge of investigative research involving the beneficial use of cannabinoids as novel interventional alternatives for multiple sclerosis (MS) and associated neuropathic pain (NPP). Using an experimental autoimmune encephalomyelitis (EAE) animal model of MS, we demonstrate the therapeutic effectiveness of two cannabinoid oil extract formulations (10:10 & 1:20 - tetrahydrocannabinol/cannabidiol) treatment. Our research findings confirm that cannabinoid treatment produces significant improvements in neurological disability scoring and behavioral assessments of NPP that directly result from their ability to reduce tumor necrosis factor alpha (TNF-α) production and enhance brain derived neurotrophic factor (BDNF) production. Henceforth, this research represents a critical step in advancing the literature by scientifically validating the merit for medical cannabinoid use and sets the foundation for future clinical trials.

DNA hydroxymethylation changes in response to spinal cord damage in a multiple sclerosis mouse model.

AIM: Roles of DNA 5-hydroxymethylcytosine (5hmC) in myelin repair were investigated in an experimental autoimmune encephalomyelitis (EAE) mouse model via its regulation on BDNF. METHODS: DNA 5hmC level and its limiting enzymes were detected in EAE mice. RESULTS: Global 5hmC modification, Tet1 and Tet2 significantly decreased in the spinal cord tissues of EAE mice. BDNF protein and mRNA decreased and were highly associated with BDNF 5hmC. Vitamin C, a Tet co-factor, increased global DNA 5hmC and reduced the neurological deficits at least by increasing BDNF 5hmC modification and protein levels. CONCLUSION: Tet protein-mediated 5hmC modifications represent a critical target involved in EAE-induced myelin damage. Targeting epigenetic modification may be a therapeutic strategy for multiple sclerosis.

Crosstalks between mTORC1 and mTORC2 variagate cytokine signaling to control NK maturation and effector function.

The metabolic checkpoint kinase mechanistic/mammalian target of rapamycin (mTOR) regulates natural killer (NK) cell development and function, but the exact underlying mechanisms remain unclear. Here, we show, via conditional deletion of Raptor (mTORC1) or Rictor (mTORC2), that mTORC1 and mTORC2 promote NK cell maturation in a cooperative and non-redundant manner, mainly by controlling the expression of Tbx21 and Eomes. Intriguingly, mTORC1 and mTORC2 regulate cytolytic function in an opposing way, exhibiting promoting and inhibitory effects on the anti-tumor ability and metabolism, respectively. mTORC1 sustains mTORC2 activity by maintaining CD122-mediated IL-15 signaling, whereas mTORC2 represses mTORC1-modulated NK cell effector functions by restraining STAT5-mediated SLC7A5 expression. These positive and negative crosstalks between mTORC1 and mTORC2 signaling thus variegate the magnitudes and kinetics of NK cell activation, and help define a paradigm for the modulation of NK maturation and effector functions.

Toll-like receptor-4/p38 MAPK signaling in the dorsal horn contributes to P2X4 receptor activation and BDNF over-secretion in cancer induced bone pain.

Our previous research suggested that the P2X4 receptor (P2X4R) expression in microglia was involved in the activation of toll-like receptor-4 (TLR4) in the dorsal horn in the rat model of cancer induced bone pain (CIBP). In this study, we focused on whether TLR4- mitogen-activated protein kinases, p38 (p38 MAPK) contributes to P2X4R activation and brain-derived neurotrophic factor (BDNF) over-secretion in CIBP. In in vitro experiment, the results showed that BDNF expression evoked by ATP stimulation was dependent on TLR4-p38. In in vivo experiment, the results demonstrated that an intrathecal injection of TLR4 siRNA alleviated nociception induced by lipopolysaccharide (LPS) plus ATP or CIBP with decreased expression of P2X4R, TLR4, BDNF, interleukin-6 (IL-6) and phosphorylated-p38 MAPK (p-p38 MAPK). Moreover, injection with p38MAPK inhibitor SB203580 resulted in an identical pattern compared with treatment with TLR4 siRNA. Our results demonstrate that the activation of TLR4-p38MAPK-P2X4R signaling in microglial possibility plays an important role in the process of nociceptive transmission in CIBP, suggesting new mechanism and potential therapeutic targets for CIBP.

Sustained elevation of NF-κB activity sensitizes offspring of maternal inflammation to hypertension via impairing PGC-1α recovery.

Growing evidence has demonstrated that maternal detrimental factors, including inflammation, contribute to the development of hypertension in the offspring. The current study found that offspring subjected to prenatal exposure of inflammation by lipopolysaccharide (LPS) challenge during the second semester showed significantly increased systolic blood pressure. In addition, these offspring also displayed augmented vascular damage and reactive oxygen species (ROS) levels in thoracic aortas when challenged with deoxycorticosterone acetate and high-salt diet (DOCA-salt). Interestingly, the antioxidant N-acetyl-L-cysteine markedly reversed these changes. Mechanistically, prenatal LPS exposure led to pre-existing elevated peroxisome proliferators-activated receptor-γ co-activator (PGC)-1α, a critical master of ROS metabolism, which up-regulated the ROS defense capacity and maintained the balance of ROS generation and elimination under resting state. However, continued elevation of NF-κB activity significantly suppressed the rapid recovery of PGC-1α expression response to DOCA-salt challenge in offspring that underwent prenatal inflammatory stimulation. This was further confirmed by using a NF-κB inhibitor (N-p-Tosyl-L-phenylalanine chloromethyl ketone) that restored PGC-1α recovery and prevented blood pressure elevation induced by DOCA-salt. Our results suggest that maternal inflammation programmed proneness to NF-κB over-activation which impaired PGC-1α-mediated anti-oxidant capacity resulting in the increased sensitivity of offspring to hypertensive damage.

Maternal inflammation activated ROS-p38 MAPK predisposes offspring to heart damages caused by isoproterenol via augmenting ROS generation.

Maternal inflammation contributes to the increased incidence of adult cardiovascular disease. The current study investigated the susceptibility of cardiac damage responding to isoproterenol (ISO) in adult offspring that underwent maternal inflammation (modeled by pregnant Sprague-Dawley rats with lipopolysaccharides (LPS) challenge). We found that 2 weeks of ISO treatment in adult offspring of LPS-treated mothers led to augmented heart damage, characterized by left-ventricular systolic dysfunction, cardiac hypertrophy and myocardial fibrosis. Mechanistically, prenatal exposure to LPS led to up-regulated expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidases, antioxidant enzymes, and p38 MAPK activity in left ventricular of adult offspring at resting state. ISO treatment exaggerated ROS generation, p38 MAPK activation but down-regulated reactive oxygen species (ROS) elimination capacity in the left ventricular of offspring from LPS-treated mothers, while antioxidant N-acetyl-L-cysteine (NAC) reversed these changes together with improved cardiac functions. The p38 inhibitor SB202190 alleviated the heart damage only via inhibiting the expression of NADPH oxidases. Collectively, our data demonstrated that prenatal inflammation programs pre-existed ROS activation in the heart tissue, which switches on the early process of oxidative damages on heart rapidly through a ROS-p38 MAPK-NADPH oxidase-ROS positive feedback loop in response to a myocardial hypertrophic challenge in adulthood.

Transcriptional Regulation of Brain-Derived Neurotrophic Factor (BDNF) by Methyl CpG Binding Protein 2 (MeCP2): a Novel Mechanism for Re-Myelination and/or Myelin Repair Involved in the Treatment of Multiple Sclerosis (MS).

Multiple sclerosis (MS) is a chronic progressive, neurological disease characterized by the targeted immune system-mediated destruction of central nervous system (CNS) myelin. Autoreactive CD4+ T helper cells have a key role in orchestrating MS-induced myelin damage. Once activated, circulating Th1-cells secrete a variety of inflammatory cytokines that foster the breakdown of blood-brain barrier (BBB) eventually infiltrating into the CNS. Inside the CNS, they become reactivated upon exposure to the myelin structural proteins and continue to produce inflammatory cytokines such as tumor necrosis factor α (TNFα) that leads to direct activation of antibodies and macrophages that are involved in the phagocytosis of myelin. Proliferating oligodendrocyte precursors (OPs) migrating to the lesion sites are capable of acute remyelination but unable to completely repair or restore the immune system-mediated myelin damage. This results in various permanent clinical neurological disabilities such as cognitive dysfunction, fatigue, bowel/bladder abnormalities, and neuropathic pain. At present, there is no cure for MS. Recent remyelination and/or myelin repair strategies have focused on the role of the neurotrophin brain-derived neurotrophic factor (BDNF) and its upstream transcriptional repressor methyl CpG binding protein (MeCP2). Research in the field of epigenetic therapeutics involving histone deacetylase (HDAC) inhibitors and lysine acetyl transferase (KAT) inhibitors is being explored to repress the detrimental effects of MeCP2. This review will address the role of MeCP2 and BDNF in remyelination and/or myelin repair and the potential of HDAC and KAT inhibitors as novel therapeutic interventions for MS.

Quetiapine Inhibits Microglial Activation by Neutralizing Abnormal STIM1-Mediated Intercellular Calcium Homeostasis and Promotes Myelin Repair in a Cuprizone-Induced Mouse Model of Demyelination.

Microglial activation has been considered as a crucial process in the pathogenesis of neuroinflammation and psychiatric disorders. Several antipsychotic drugs (APDs) have been shown to display inhibitory effects on microglial activation in vitro, possibly through the suppression of elevated intracellular calcium (Ca(2+)) concentration. However, the exact underlying mechanisms still remain elusive. In this study, we aimed to investigate the inhibitory effects of quetiapine (Que), an atypical APD, on microglial activation. We utilized a chronic cuprizone (CPZ)-induced demyelination mouse model to determine the direct effect of Que on microglial activation. Our results showed that treatment with Que significantly reduced recruitment and activation of microglia/macrophage in the lesion of corpus callosum and promoted remyelination after CPZ withdrawal. Our in vitro studies also confirmed the direct effect of Que on lipopolysaccharide (LPS)-induced activation of microglial N9 cells, whereby Que significantly inhibited the release of nitric oxide (NO) and tumor necrosis factor α (TNF-α). Moreover, we demonstrated that pretreatment with Que, neutralized the up-regulation of STIM1 induced by LPS and declined both LPS and thapsigargin (Tg)-induced store-operated Ca(2+) entry (SOCE). Finally, we found that pretreatment with Que significantly reduced the translocation of nuclear factor kappa B (NF-κB) p65 subunit from cytoplasm to nuclei in LPS-activated primary microglial cells. Overall, our data suggested that Que may inhibit microglial activation by neutralization of the LPS-induced abnormal STIM1-mediated intercellular calcium homeostasis.

Metabolic disposition of the insect repellent DEET and the sunscreen oxybenzone following intravenous and skin administration in rats.

Insect repellent N,N-diethyl-m-toluamide (DEET) and sunscreen oxybenzone have shown a synergistic percutaneous enhancement when applied concurrently. Both compounds are extensively metabolized in vivo into a series of potentially toxic metabolites: 2 metabolites of DEET, N,N-diethyl-m-hydroxymethylbenzamide (DHMB) and N-ethyl-m-toluamide (ET), and 3 metabolites of oxybenzone, 2,4-dihydroxybenzophenone (DHB), 2,2-dihydroxy-4-methoxybenzophenone (DMB), and 2,3,4-trihydroxybenzophenone (THB). In this study, the metabolites were extensively distributed following intravenous and topical skin administration of DEET and oxybenzone in rats. Combined application enhanced the disposition of all DEET metabolites in the liver but did not consistently affect the distribution of oxybenzone metabolites. The DHMB appeared to be the major metabolite for DEET, while THB and its precursor DHB were the main metabolites for oxybenzone. Repeated once-daily topical application for 30 days led to higher concentrations of DEET metabolites in the liver. Hepatoma cell studies revealed a decrease in cellular proliferation from all metabolites as single and combined treatments, most notably at 72 hours. Increased accumulation of DHMB and ET in the liver together with an ability to reduce cellular proliferation at achievable plasma concentrations indicated that simultaneous exposure to DEET and oxybenzone might have the potential to precipitate adverse effects in a rat animal model.

Tissue disposition of the insect repellent DEET and the sunscreen oxybenzone following intravenous and topical administration in rats.

The insect repellent N,N-diethyl-m-toluamide (DEET) and sunscreen oxybenzone (OBZ) have been shown to produce synergistic permeation enhancement when applied concurrently in vitro and in vivo. The disposition of both compounds following intravenous administration (2 mg/kg of DEET or OBZ) and topical skin application (100 mg/kg of DEET and 40 mg/kg of OBZ) was determined in male Sprague-Dawley rats. Pharmacokinetic analysis was also conducted using compartmental and non-compartmental methods. A two-compartment model was deemed the best fit for intravenous administration. The DEET and oxybenzone permeated across the skin to accumulate in blood, liver and kidney following topical skin application. Combined use of DEET and oxybenzone accelerated the disappearance of both compounds from the application site, increased their distribution in the liver and significantly decreased the apparent elimination half-lives of both compounds (p < 0.05). Hepatoma cell studies revealed toxicity from exposure to all treatment concentrations, most notably at 72 h. Although DEET and oxybenzone were capable of mutually enhancing their percutaneous permeation and systemic distribution from topical skin application, there was no evidence of increased hepatotoxic deficits from concurrent application.

Axotomy-induced up-regulation of tumor necrosis factor-alpha in the dorsal root ganglia.

OBJECTIVES: Neuropathic pain is a chronic pain syndrome associated with drug, injury or disease-induced damage or destruction of sensory afferent fibers of the dorsal root ganglia (DRG). Although the exact underlying pathologic mechanisms are not known, pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) are recognized as potential modulators of peripheral and central nervous system inflammatory responses. They play a crucial role in injury and the pathologic development of chronic pain syndromes such as neuropathic pain. METHODS: Twenty-four rats were divided into a naive control (n=6), sham (surgery exposing sciatic nerve, n=6), and peripheral nerve lesion group (unilateral axotomy of sciatic nerve, n=12). RESULTS: The results of this study demonstrate a transient up-regulation of TNF-alpha expression within ipsi- and contralateral DRG following complete unilateral sciatic nerve axotomy as confirmed by immunohistochemistry, reverse transcriptase-polymerase chain reaction (RT-PCR) and real-time PCR. Elevated expression of TNF-alpha was noted to occur within the first 7 days post-axotomy, which subsequently normalized to baseline levels by day 14. This transient up-regulation was also associated with a switch in cellular source from predominant satellite cell expression at baseline to that involving satellite cells and abundant numbers of sensory neurons. DISCUSSION: These results support the role of TNF-alpha in the upstream cascade of cellular events involved in the underlying pathogenesis of neuropathic pain. Strategies targeting the early attenuation of TNF-alpha within the DRG during the first week post-injury may have significant clinical impact in preventing the downstream cascade of events involved in the underlying cellular pathology of neuropathic pain.