Post-surgery fluids promote transition of cancer stem cell- to-endothelial and AKT/mTOR activity, contributing to relapse of giant cell tumors of bone

Giant cell tumors of bone (GCTB) are rare sarcomas with a high rate of unpredictable local relapse. Studies suggest that surgical methods affect recurrence, supporting the idea that local disease develops from re-growth of residual cancer cells. To identify early prognostic markers of individual risk of recurrence, we evaluated the effect of post-surgery fluids from a cohort of GCTB patients on growth of primary and established sarcoma cell lines, and mice xenographts. Post-surgery fluids increased cell growth and enhanced expression of CD44++, the principal receptor for the extracellular matrix component hyaluronan and the mesenchymal stem marker CD117+. Cancer cells became highly invasive and tumorigenic, acquiring stemness properties, and activated AKT/mTOR pathway. Prolonged stimulation with post-surgery fluids down-regulated the mesenchymal gene TWIST1 and Vimentin protein, and transdifferentiated cells into tubule-like structures positive to the endothelial markers VE-Cadherin and CD31+. In mice, post-surgery fluids gave rise to larger and more vascularized tumors than control, while in patients AKT/mTOR pathway activation was associated with recurrence by logistic regression (Kaplan-Meier; P<0.001). These findings indicate that post-surgery fluids are an adjuvant in mechanisms of tumor regrowth, increasing stem cell growth and AKT/mTOR activity

Phytocannabinoids promote viability and functional adipogenesis of bone marrow-derived mesenchymal stem cells through different molecular targets.

Abstract The cellular microenvironment plays a critical role in the maintenance of bone marrow-derived mesenchymal stem cells (BM-MSCs) and their subsequent cell lineage differentiation. Recent studies suggested that individuals with adipocyte-related metabolic disorders have altered function and adipogenic potential of adipose stem cell subpopulations, primarily BM-MSCs, increasing the risk of heart attack, stroke or diabetes. In this study, we explored the potential therapeutic effect of some of the most abundant non-euphoric compounds derived from the Cannabis sativa plant (or phytocannabinoids) including tetrahydrocannabivarin (THCV), cannabidiol (CBD), cannabigerol (CBG), cannabidiolic acid (CBDA) and cannabigerolic acid (CBGA), by analysing their pharmacological activity on viability of endogenous BM-MSCs as well as their ability to alter BM-MSC proliferation and differentiation into mature adipocytes. We provide evidence that CBD, CBDA, CBGA and THCV (5 µM) increase the number of viable BM-MSCs; whereas only CBG (5 µM) and CBD (5 µM) alone or in combination promote BM-MSCs maturation into adipocytes via distinct molecular mechanisms. These effects were revealed both in vitro and in vivo. In addition, phytocannabinoids prevented the insulin signalling impairment induced by palmitate in adipocytes differentiated from BM-MSCs. Our study highlights phytocannabinoids as a potential novel pharmacological tool to regain control of functional adipose tissue in unregulated energy homeostasis often occurring in metabolic disorders including type 2 diabetes mellitus (T2DM), aging and lipodystrophy

EP1 receptor within the ventrolateral periaqueductal grey controls thermonociception and rostral ventromedial medulla cell activity in healthy and neuropathic rat

The aim of this study was to investigate the expression of prostaglandin EP1 receptor within the ventrolateral periaqueductal grey (VL PAG). The role of VL PAG EP1 receptor in controlling thermonociception and rostral ventromedial medulla (RVM) activity in healthy and neuropathic rats was also examined. EP1 receptor was indeed found to be expressed within the VL PAG and co-localized with vesicular GABA transporter. Intra-VL PAG microinjection of ONO-DI-004, a selective EP1 receptor agonist, dose-dependently reduced tail flick latency as well as respectively increasing and decreasing the spontaneous activity of ON and OFF cells. Furthermore, it increased the ON cell burst and OFF cell pause. Intra-VL PAG prostaglandin E2 (PGE2) behaved similarly to ONO-DI-004. The effects of ONO-DI-004 and PGE2 were antagonized by intra-VL PAG L335677, a selective EP1 receptor antagonist. L335677 dose-dependently increased the tail flick latency and ongoing activity of the OFF cells, while reducing the ongoing ON cell activity. It also decreased the ON cell burst and OFF cell pause. In neuropathic rats using spare nerve injury (SNI) of the sciatic nerve model, EP1 receptor expression decreased in the VL PAG. However, ONO-DI-004 and L335677 were able to alter pain responses and ON and OFF cell activity, as they did in healthy animals. Collectively, these data show that within the VL PAG, EP1 receptor has a facilitatory effect on the nociceptive response and consistently affects RVM neuron activity. Thus, the blockade of EP1 receptor in the VL PAG leads to antinociception in neuropathic pain conditions, despite its down-regulation. The expression of EP1 receptor on GABAergic neurons is consistent with an EP1 receptor blockade-induced disinhibition of the antinociceptive descending pathway at VL PAG level

Altered gut microbiota and endocannabinoid system tone in vitamin D deficiency-mediated chronic pain

Abstract Recent evidence points to the gut microbiota as a regulator of brain and behavior, although it remains to be determined if gut bacteria play a role in chronic pain. The endocannabinoid system is implicated in inflammation and chronic pain processing at both the gut and central nervous system (CNS) levels. In the present study, we used low Vitamin D dietary intake in mice and evaluated possible changes in gut microbiota, pain processing and endocannabinoid system signaling. Vitamin D deficiency induced a lower microbial diversity characterized by an increase in Firmicutes and a decrease in Verrucomicrobia and Bacteroidetes. Concurrently, vitamin D deficient mice showed tactile allodynia associated with neuronal hyperexcitability and alterations of endocannabinoid system members (endogenous mediators and their receptors) at the spinal cord level. Changes in endocannabinoid (anandamide and 2-arachidonoylglycerol) levels were also observed in the duodenum and colon. Remarkably, the anti-inflammatory anandamide congener, palmitoylethanolamide, counteracted both the pain behaviour and spinal biochemical changes in vitamin D deficient mice, whilst increasing the levels of Akkermansia, Eubacterium and Enterobacteriaceae, as compared with vehicle-treated mice. Finally, induction of spared nerve injury in normal or vitamin D deficient mice was not accompanied by changes in gut microbiota composition. Our data suggest the existence of a link between Vitamin D deficiency – with related changes in gut bacterial composition – and altered nociception, possibly via molecular mechanisms involving the endocannabinoid and related mediator signaling systems

2-Pentadecyl-2-oxazoline ameliorates memory impairment and depression-like behaviour in neuropathic mice: possible role of adrenergic alpha2- and H3 histamine autoreceptors

Neuropathic pain (NP) remains an untreatable disease due to the complex pathophysiology that involves the whole pain neuraxis including the forebrain. Sensory dysfunctions such as allodynia and hyperalgesia are only part of the symptoms associated with neuropathic pain that extend to memory and affectivity deficits. The development of multi-target molecules might be a promising therapeutic strategy against the symptoms associated with NP. 2-pentadecyl-2-oxazoline (PEA-OXA) is a plant-derived agent, which has shown effectiveness against chronic pain and associated neuropsychiatric disorders. The molecular mechanisms by which PEA-OXA exerts its effects are, however, only partially known. In the current study, we show that PEA-OXA, besides being an alpha2 adrenergic receptor antagonist, also acts as a modulator at histamine H3 receptors, and report data on its effects on sensory, affective and cognitive symptoms associated with the spared nerve injury (SNI) model of neuropathic pain in mice. Treatment for 14 days with PEA-OXA after the onset of the symptoms associated with neuropathic pain resulted in the following effects: (i) allodynia was decreased; (ii) affective/cognitive impairment associated with SNI (depression, spatial, and working memories) was counteracted; (iii) long-term potentiation in vivo in the lateral entorhinal cortex-dentate gyrus (perforant pathway, LPP) was ameliorated, (iv) hippocampal glutamate, GABA, histamine, norepinephrine and dopamine level alterations after peripheral nerve injury were reversed, (v) expression level of the TH positive neurons in the Locus Coeruleus were normalized. Thus, a 16-day treatment with PEA-OXA alleviates the sensory, emotional, cognitive, electrophysiological and neurochemical alterations associated with SNI-induced neuropathic pain

Oral Cannabidiol Prevents Allodynia and Neurological Dysfunctions in a Mouse Model of Mild Traumatic Brain Injury

Neurological dysfunctions are the most impactful and persistent consequences of traumatic brain injury (TBI). Indeed, previous reports suggest that an association between TBI and chronic pain syndromes, as well anxio-depressive behaviors, tends to be more common in patients with mild forms of TBI. At present, no effective treatment options are available for these symptoms. In the present study, we used a weight drop mild TBI mouse model to investigate the effect of a commercially available 10% Cannabidiol (CBD) oil on both the sensorial and neuropsychiatric dysfunctions associated with mild TBI through behavioral and biomolecular approaches. TBI mice developed chronic pain associated with anxious and aggressive behavior, followed by a late depressive-like behavior and impaired social interaction. Such behaviors were related with specific changes in neurotransmitters release at cortical levels. CBD oral treatment restored the behavioral alterations and partially normalized the cortical biochemical changes. In conclusion, our data show some of the brain modifications probably responsible for the behavioral phenotype associated with TBI and suggest the CBD as a pharmacological tool to improve neurological dysfunctions caused by the trauma

The blockade of the transient receptor potential vanilloid type 1 and fatty acid amide hydrolase decreases symptoms and central sequelae in the medial prefrontal cortex of neuropathic rats

<p>Abstract</p> <p>Background</p> <p>Neuropathic pain is a chronic disease resulting from dysfunction within the "pain matrix". The basolateral amygdala (BLA) can modulate cortical functions and interactions between this structure and the medial prefrontal cortex (mPFC) are important for integrating emotionally salient information. In this study, we have investigated the involvement of the transient receptor potential vanilloid type 1 (TRPV1) and the catabolic enzyme fatty acid amide hydrolase (FAAH) in the morphofunctional changes occurring in the pre-limbic/infra-limbic (PL/IL) cortex in neuropathic rats.</p> <p>Results</p> <p>The effect of <it>N</it>-arachidonoyl-serotonin (AA-5-HT), a hybrid FAAH inhibitor and TPRV1 channel antagonist, was tested on nociceptive behaviour associated with neuropathic pain as well as on some phenotypic changes occurring on PL/IL cortex pyramidal neurons. Those neurons were identified as belonging to the BLA-mPFC pathway by electrical stimulation of the BLA followed by hind-paw pressoceptive stimulus application. Changes in their spontaneous and evoked activity were studied in sham or spared nerve injury (SNI) rats before or after repeated treatment with AA-5-HT. Consistently with the SNI-induced changes in PL/IL cortex neurons which underwent profound phenotypic reorganization, suggesting a profound imbalance between excitatory and inhibitory responses in the mPFC neurons, we found an increase in extracellular glutamate levels, as well as the up-regulation of FAAH and TRPV1 in the PL/IL cortex of SNI rats. Daily treatment with AA-5-HT restored cortical neuronal activity, normalizing the electrophysiological changes associated with the peripheral injury of the sciatic nerve. Finally, a single acute intra-PL/IL cortex microinjection of AA-5-HT transiently decreased allodynia more effectively than URB597 or I-RTX, a selective FAAH inhibitor or a TRPV1 blocker, respectively.</p> <p>Conclusion</p> <p>These data suggest a possible involvement of endovanilloids in the cortical plastic changes associated with peripheral nerve injury and indicate that therapies able to normalize endovanilloid transmission may prove useful in ameliorating the symptoms and central sequelae associated with neuropathic pain.</p

Discovery of Prostamide F2α and Its Role in Inflammatory Pain and Dorsal Horn Nociceptive Neuron Hyperexcitability

It was suggested that endocannabinoids are metabolized by cyclooxygenase (COX)-2 in the spinal cord of rats with kaolin/λ-carrageenan-induced knee inflammation, and that this mechanism contributes to the analgesic effects of COX-2 inhibitors in this experimental model. We report the development of a specific method for the identification of endocannabinoid COX-2 metabolites, its application to measure the levels of these compounds in tissues, and the finding of prostamide F2α (PMF2α) in mice with knee inflammation. Whereas the levels of spinal endocannabinoids were not significantly altered by kaolin/λ-carrageenan-induced knee inflammation, those of the COX-2 metabolite of AEA, PMF2α, were strongly elevated. The formation of PMF2α was reduced by indomethacin (a non-selective COX inhibitor), NS-398 (a selective COX-2 inhibitor) and SC-560 (a selective COX-1 inhibitor). In healthy mice, spinal application of PMF2α increased the firing of nociceptive (NS) neurons, and correspondingly reduced the threshold of paw withdrawal latency (PWL). These effects were attenuated by the PMF2α receptor antagonist AGN211336, but not by the FP receptor antagonist AL8810. Also prostaglandin F2α increased NS neuron firing and reduced the threshold of PWL in healthy mice, and these effects were antagonized by AL8810, and not by AGN211336. In mice with kaolin/λ-carrageenan-induced knee inflammation, AGN211336, but not AL8810, reduced the inflammation-induced NS neuron firing and reduction of PWL. These findings suggest that inflammation-induced, and prostanoid-mediated, enhancement of dorsal horn NS neuron firing stimulates the production of spinal PMF2α, which in turn contributes to further NS neuron firing and pain transmission by activating specific receptors