Teresa Iuvone

Involvement of NF‐κB in the regulation of cyclooxygenase‐2 protein expression in LPS‐stimulated J774 macrophages

We investigated the involvement of NF‐κB in the regulation of COX‐2 protein expression and prostaglandin production in LPS‐stimulated J774 macrophages. Incubation of J774 cells with LPS (1 μg/ml) for 24 h caused an increase of COX‐2 protein expression and accumulation of both PGE2 and 6‐keto‐PGF1α in the cell culture medium. Ammonium pyrrolidinedithiocarbamate (APDC, 0.1, 1, 10 μM) and N‐α‐p‐tosyl‐l‐lysine chloromethylketone (TLCK, 1, 10, 100 μM), two inhibitors of NF‐κB activation, suppressed in a concentration‐dependent manner both LPS‐induced COX‐2 protein expression and prostanoid generation. Moreover, APDC and TLCK both inhibited the LPS‐induced increase of NF‐κB DNA binding activity and prevented IκB‐α degradation. Our results show for the first time that NF‐κB is involved in COX‐2 protein expression in LPS‐stimulated J774 macrophages and suggest that inhibitors of NF‐κB activation may represent a useful tool for the pharmacological control of inflammation.

Cannabinoid CB1‐receptor mediated regulation of gastrointestinal motility in mice in a model of intestinal inflammation

We have studied the effect of cannabinoid agonists (CP 55,940 and cannabinol) on intestinal motility in a model of intestinal inflammation (induced by oral croton oil in mice) and measured cannabinoid receptor expression, endocannabinoids (anandamide and 2‐arachidonylglycerol) and anandamide amidohydrolase activity both in physiological and pathophysiological states. CP 55,940 (0.03 – 10 nmol mouse−1) and cannabinol (10 – 3000 nmol mouse−1) were more active in delaying intestinal motility in croton oil‐treated mice than in control mice. These inhibitory effects were counteracted by the selective cannabinoid CB1 receptor antagonist SR141716A (16 nmol mouse−1). SR141716A (1 – 300 nmol mouse−1), administered alone, increased intestinal motility to the same extent in both control and croton oil‐treated mice Croton oil‐induced intestinal inflammation was associated with an increased expression of CB1 receptor, an unprecedented example of up‐regulation of cannabinoid receptors during inflammation. High levels of anandamide and 2‐arachidonylglycerol were detected in the small intestine, although no differences were observed between control and croton oil‐treated mice; by contrast anandamide amidohydrolase activity increased 2 fold in the inflamed small intestine. It is concluded that inflammation of the gut increases the potency of cannabinoid agonists possibly by ‘up‐regulating’ CB1 receptor expression; in addition, endocannabinoids, whose turnover is increased in inflamed gut, might tonically inhibit intestinal motility.

Neuroprotective effect of cannabidiol, a non-psychoactive component from Cannabis sativa, on β-amyloid-induced toxicity in PC12 cells

Alzheimers disease is widely held to be associated with oxidative stress due, in part, to the membrane action of β‐amyloid peptide aggregates. Here, we studied the effect of cannabidiol, a major non‐psychoactive component of the marijuana plant (Cannabis sativa) on β‐amyloid peptide‐induced toxicity in cultured rat pheocromocytoma PC12 cells. Following exposure of cells to β‐amyloid peptide (1 µg/mL), a marked reduction in cell survival was observed. This effect was associated with increased reactive oxygen species (ROS) production and lipid peroxidation, as well as caspase 3 (a key enzyme in the apoptosis cell‐signalling cascade) appearance, DNA fragmentation and increased intracellular calcium. Treatment of the cells with cannabidiol (10−7−10−4m) prior to β‐amyloid peptide exposure significantly elevated cell survival while it decreased ROS production, lipid peroxidation, caspase 3 levels, DNA fragmentation and intracellular calcium. Our results indicate that cannabidiol exerts a combination of neuroprotective, anti‐oxidative and anti‐apoptotic effects against β‐amyloid peptide toxicity, and that inhibition of caspase 3 appearance from its inactive precursor, pro‐caspase 3, by cannabidiol is involved in the signalling pathway for this neuroprotection.

The spice sage and its active ingredient rosmarinic acid protect PC12 cells from amyloid-β peptide-induced neurotoxicity

Traditional use and clinical reports suggest that the culinary herb sage (Salvia officinalis) may be effective for patients with mild to moderate Alzheimers disease (AD). In this study, we evaluated the effect of a standardized extract from the leaves of S. officinalis (SOE) and its active ingredient rosmarinic acid on Alzheimer amyloid-β peptide (Aβ)-induced toxicity in cultured rat pheochromocytoma (PC12) cells. Incubation of PC12 cells with Aβ (fragment 1–42) for 24 h caused cell death, and this effect was reduced by SOE and its active ingredient, rosmarinic acid. Rosmarinic acid reduced a number of events induced by Aβ. These include reactive oxygen species formation, lipid peroxidation, DNA fragmentation, caspase-3 activation, and tau protein hyperphosphorylation. Moreover, rosmarinic acid inhibited phosphorylated p38 mitogen-activated protein kinase but not glycogen synthase kinase 3β activation. These data show the neuroprotective effect of sage against Aβ-induced toxicity, which could validate the traditional use of this spice in the treatment of AD. Rosmarinic acid could contribute, at least in part, for sage-induced neuroprotective effect.

Cannabidiol In vivo blunts β-amyloid induced neuroinflammation by suppressing IL-1β and iNOS expression

Pharmacological inhibition of beta‐amyloid (Aβ) induced reactive gliosis may represent a novel rationale to develop drugs able to blunt neuronal damage and slow the course of Alzheimers disease (AD). Cannabidiol (CBD), the main non‐psychotropic natural cannabinoid, exerts in vitro a combination of neuroprotective effects in different models of Aβ neurotoxicity. The present study, performed in a mouse model of AD‐related neuroinflammation, was aimed at confirming in vivo the previously reported antiinflammatory properties of CBD.

Cannabidiol inhibits inducible nitric oxide synthase protein expression and nitric oxide production in β-amyloid stimulated PC12 neurons through p38 MAP kinase and NF-κB involvement

In view of the pro-inflammatory scenario observed in Alzheimers disease, in the recent years anti-inflammatory drugs have been proposed as potential therapeutic agents. We have previously shown that cannabidiol, the main non-psychotropic component from Cannabis sativa, possess a variegate combination of anti-oxidant and anti-apoptotic effects that protect PC12 cells from Abeta toxicity. In parallel, cannabidiol has been described to have anti-inflammatory properties in acute models of inflammation but the possible inhibitory effect of cannabidiol on iNOS protein expression and nitrite production in the nitrosative stress induced by Abeta in neuronal cell-line is un-investigated. Stimulation of differentiated PC12 cells with Abeta (1-42) (1 microg/mL) for 36 h caused a significant increase of nitrite production, compared to un-stimulated cells, that was inhibited in a concentration-dependent manner by both the non-selective iNOS inhibitor, L-NAME (0.3-30 microM), and, at higher extent, by the selective iNOS inhibitor SMT (0.3-30 microM). CBD (10(-6) to 10(-4) M) inhibited both nitrite production and iNOS protein expression induced by Abeta (1-42). Cannabidiol effect was mediated through the inhibition of phosphorylated form of p38 MAP kinase and transcription factor nuclear factor-kappaB activation in a concentration-dependent manner. The here reported data increases our knowledge about the possible neuroprotective mechanism of cannabidiol, highlighting the importance of this compound to inhibit beta-amyloid induced neurodegeneration, in view of its low toxicity in humans.

The marijuana component cannabidiol inhibits β-amyloid-induced tau protein hyperphosphorylation through Wnt/β-catenin pathway rescue in PC12 cells

Alzheimer’s disease (AD) is the most common age-related neurodegenerative disorder. A massive accumulation of β-amyloid (Aβ) peptide aggregates has been proposed as pivotal event in AD. Aβ-induced toxicity is accompanied by a variegated combination of events including oxidative stress. The Wnt pathway has multiple actions in the cascade of events triggered by Aβ, and drugs that rescue Wnt activity may be considered as novel therapeutics for AD treatment. Cannabidiol, a non-psychoactive marijuana component, has been recently proposed as an antioxidant neuroprotective agent in neurodegenerative diseases. Moreover, it has been shown to rescue PC12 cells from toxicity induced by Aβ peptide. However, the molecular mechanism of cannabidiol-induced neuroprotective effect is still unknown. Here, we report that cannabidiol inhibits hyperphosphorylation of tau protein in Aβ-stimulated PC12 neuronal cells, which is one of the most representative hallmarks in AD. The effect of cannabidiol is mediated through the Wnt/β-catenin pathway rescue in Aβ-stimulated PC12 cells. These results provide new molecular insight regarding the neuroprotective effect of cannabidiol and suggest its possible role in the pharmacological management of AD, especially in view of its low toxicity in humans.

Cannabidiol: A Promising Drug for Neurodegenerative Disorders?

Neurodegenerative diseases represent, nowadays, one of the main causes of death in the industrialized country. They are characterized by a loss of neurons in particular regions of the nervous system. It is believed that this nerve cell loss underlies the subsequent decline in cognitive and motor function that patients experience in these diseases. A range of mutant genes and environmental toxins have been implicated in the cause of neurodegenerative disorders but the mechanism remains largely unknown. At present, inflammation, a common denominator among the diverse list of neurodegenerative diseases, has been implicated as a critical mechanism that is responsible for the progressive nature of neurodegeneration. Since, at present, there are few therapies for the wide range of neurodegenerative diseases, scientists are still in search of new therapeutic approaches to the problem. An early contribution of neuroprotective and antiinflammatory strategies for these disorders seems particularly desirable because isolated treatments cannot be effective. In this contest, marijuana derivatives have attracted special interest, although these compounds have always raised several practical and ethical problems for their potential abuse. Nevertheless, among Cannabis compounds, cannabidiol (CBD), which lacks any unwanted psychotropic effect, may represent a very promising agent with the highest prospect for therapeutic use.

N-palmitoyl-ethanolamine: Biochemistry and new therapeutic opportunities.

Although its presence in mammalian tissues has been known since the 1960s, N-palmitoyl-ethanolamine (PEA) has emerged only recently among other bioactive N-acylethanolamines as an important local pro-homeostatic mediator which, due to its chemical stability, can be also administered exogenously as the active principle of current anti-inflammatory and analgesic preparations (e.g. Normast, Pelvilen). Much progress has been made towards the understanding of the mechanisms regulating both the tissue levels of PEA under physiological and pathological conditions, and its pharmacological actions. Here we review these new developments in PEA biochemistry and pharmacology, and discuss novel potential indications for the therapeutic use of this compound and of synthetic tools that selectively retard its catabolism, such as the inhibitors of the recently cloned N-acylethanolamine-hydrolyzing acid amidase.

Cannabidiol reduces Aβ-induced neuroinflammation and promotes hippocampal neurogenesis through PPARγ involvement

Peroxisome proliferator-activated receptor-γ (PPARγ) has been reported to be involved in the etiology of pathological features of Alzheimers disease (AD). Cannabidiol (CBD), a Cannabis derivative devoid of psychomimetic effects, has attracted much attention because of its promising neuroprotective properties in rat AD models, even though the mechanism responsible for such actions remains unknown. This study was aimed at exploring whether CBD effects could be subordinate to its activity at PPARγ, which has been recently indicated as its putative binding site. CBD actions on β-amyloid-induced neurotoxicity in rat AD models, either in presence or absence of PPAR antagonists were investigated. Results showed that the blockade of PPARγ was able to significantly blunt CBD effects on reactive gliosis and subsequently on neuronal damage. Moreover, due to its interaction at PPARγ, CBD was observed to stimulate hippocampal neurogenesis. All these findings report the inescapable role of this receptor in mediating CBD actions, here reported.