Search

Purpose: This study elucidates the mechanism of the physiological effect of cannabidiol (CBD) by assessing its impact on lipopolysaccharide (LPS)-induced inflammation in RWPE-1 cells and prostatitis-induced by 17β-estradiol and dihydrotestosterone in a rat model, focusing on its therapeutic potential for chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS). Materials and Methods: RWPE-1 cells were stratified in vitro into three groups: (1) controls, (2) cells with LPS-induced inflammation, and (3) cells with LPS-induced inflammation and treated with CBD. Enzyme-linked immunosorbent assays and western blots were performed on cellular components and supernatants after administration of CBD. Five groups of six Sprague– Dawley male rats were assigned: (1) control, (2) CP/CPPS, (3) CP/CPPS and treated with 50 mg/kg CBD, (4) CP/CPPS and treated with 100 mg/kg CBD, and (5) CP/CPPS and treated with 150 mg/kg CBD. Prostatitis was induced through administration of 17β-estradiol and dihydrotestosterone. After four weeks of CBD treatment, a pain index was evaluated, and prostate tissue was collected for subsequent histologic examination and western blot analysis. Results: CBD demonstrated efficacy in vivo for CP/CPPS and in vitro for inflammation. It inhibited the toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB) pathway by activating the CB2 receptor, reducing expression of interleukin-6, tumor necrosis factor-alpha, and cyclooxygenase-2 (COX2) (p

Abstract To test the hypothesis that genetic and pharmacological modulation of the classical cannabinoid type 1 (CB1) and 2 (CB2) receptors attenuate cancer-induced bone pain, we searched Medline, Web of Science and Scopus for relevant skeletal and non-skeletal cancer studies from inception to July 28, 2022. We identified 29 animal and 35 human studies. In mice, a meta-analysis of pooled studies showed that treatment of osteolysis-bearing males with the endocannabinoids AEA and 2-AG (mean difference [MD] − 24.83, 95% confidence interval [95%CI] − 34.89, − 14.76, p

Abstract Introduction The cannabinoid receptor 2 (CB2) is mainly involved in the immune system. However, although CB2 has been reported to play an anti‐tumor function in breast cancer (BC), its specific mechanism in BC remains unclear. Methods We examined the expression and prognostic significance of CB2 in BC tissues by qPCR, second‐generation sequencing, western blot, and immunohistochemistry. We assessed the impacts of overexpression and a specific agonist of CB2 on the growth, proliferation, apoptosis, and drug resistance of BC cells in vitro and in vivo using CCK‐8, flow cytometry, TUNEL staining, immunofluorescence, tumor xenografts, western blot, and colony formation assays. Results CB2 expression was significantly lower in BC compared with paracancerous tissues. It was also highly expressed in benign tumors and ductal carcinoma in situ, and its expression was correlated with prognosis in BC patients. CB2 overexpression and treatment of BC cells with a CB2 agonist inhibited proliferation and promoted apoptosis, and these actions were achieved by suppressing the PI3K/Akt/mTOR signaling pathway. Moreover, CB2 expression was increased in MDA‐MB‐231 cell treated with cisplatin, doxorubicin, and docetaxel, and sensitivity to these anti‐tumor drugs was increased in BC cells overexpressing CB2. Conclusions These findings reveal that CB2 mediates BC via the PI3K/Akt/mTOR signaling pathway. CB2 could be a novel target for the diagnosis and treatment of BC.

Cannabinoid ligands are known to possess neuroprotective actions and may have utility in the treatment of neurodegeneration. The major targets for cannabinoids include the classical CB1 cannabinoid receptor, as well as the novel cannabinoid receptor GPR55 which binds to many synthetic cannabinoid ligands. In this study, novel thiopyranothiazoles 1, 3, 4, 6, and 7 were synthesized and their pharmacological activity as potential cannabinoid-like ligands was evaluated in glioblastoma cells, cultured cortical neurons, and cells of HEK293 line expressing GPR55. Stimulation of protein kinase ERK1/2, MAP-kinases and cAMP response element binding protein (CREB) was evaluated using Western-blot analysis. CREB activation was additionally monitored by means of confocal imaging of nuclear phospho-CREB labeling. Docking simulation confirmed the good affinity of the synthesized compounds to CB1 and CB2 receptors. Striking effects of the chromeno[4′,3′:4,5]thiopyrano[2,3-d][1,3]thiazol with ethylacetate moiety (3) and isothiochromeno[4a,4-d]thiazole with phenazone fragment (7) on pCREB activation as the indicator of stimulation of the pathway beneficial for neurons survival were observed.

Introducción: los cannabinoides son derivados de la planta de cannabis, y han captado la atención por sus efectos terapéuticos demostrados como antiinflamatorios, analgésicos, antieméticos y antitumorales. Su potencial radica en su interacción con los receptores endocannabinoides CB1 y CB2, confirmada por estudios in vitro e in vivo. Por ello, resulta interesante estudiar dicha interacción proteína-ligando mediante técnicas de acoplamiento molecular. Estos permiten describir las zonas de los cannabinoides responsables de su actividad biológica, posibilitando que estos compuestos den respuesta a patologías relacionadas con el sistema inmune. Objetivo: estudiar las interacciones proteína-ligando de 30 cannabinoides análogos del receptor CBN II. Métodos: el grupo de moléculas estudiadas en esta investigación incluyó cannabinoides endógenos, fitocannabinoides y cannabinoides sintéticos, con 30 análogos extraídos de PubChem. Se seleccionó la estructura 3D del receptor CB2 de Protein Data Bank con código 5ZTY. Se optimizó el receptor y ligandos para el acoplamiento molecular, validado con un RMSD de 1.76. Resultados: los ligandos SR141716, AM251 y JZL184 mostraron mejor afinidad por CB2. La fenilalanina fue el aminoácido más presente en la unión, con interacciones alquil y pi-alquil, sugiriendo la importancia de anillos aromáticos. Esto concuerda con otros estudios que indican que los grupos cetona en posición 3 del indol y grupos hidroxilo/metoxilo en posición 6 influyen en la afinidad de unión a CB2 mediante donación de hidrógeno al carbonilo del ligando. Conclusión: los análogos SR141716, AM251 y JZL184, relacionados con el THC y el CBD, se perfilan como moléculas prometedoras para evaluación experimental debido a su alta afinidad por el receptor CB2. Estos compuestos podrían presentar efectos beneficiosos mediados por CB2 en el tratamiento del dolor, inflamación, cáncer y trastornos inmunológicos.

Early adversity, the loss of the inhibitory GABAergic interneuron parvalbumin, and elevated neuroinflammation are associated with depression. Individuals with a maltreatment history initiate medicinal cannabis use earlier in life than non-maltreated individuals, suggesting self-medication. Female rats underwent maternal separation (MS) between 2 and 20 days of age to model early adversity or served as colony controls. The prelimbic cortex and behavior were examined to determine whether MS alters the cannabinoid receptor 2 (CB2), which has anti-inflammatory properties. A reduction in the CB2-associated regulatory enzyme MARCH7 leading to increased NLRP3 was observed with Western immunoblots in MS females. Immunohistochemistry with stereology quantified numbers of parvalbumin-immunoreactive cells and CB2 at 25, 40, and 100 days of age, revealing that the CB2 receptor associated with PV neurons initially increases at P25 and subsequently decreases by P40 in MS animals, with no change in controls. Confocal and triple-label microscopy suggest colocalization of these CB2 receptors to microglia wrapped around the parvalbumin neuron. Depressive-like behavior in MS animals was elevated at P40 and reduced with the CB2 agonist HU-308 or a CB2-overexpressing lentivirus microinjected into the prelimbic cortex. These results suggest that increasing CB2 expression by P40 in the prelimbic cortex prevents depressive behavior in MS female rats.

Abstract Background As effective immune modulators, Endocannabinoids may suppress the inflammatory responses in periodontitis. This study assessed the expression of cannabinoid receptors in gingiva and the impact on periodontitis. Methods A cross‐sectional study on 20 patients with more than stage II and Grade A periodontitis and a control group consisting of 19 healthy individuals was performed. The gingival biopsies were assessed for the expression of CB1 and CB2 using the quantitative reverse transcription polymerase chain reaction, TaqMan method. Results The study sample consisted of 39 subjects, 31 females (79.5%) and 8 males (20.5%), including 20 periodontitis subjects (80% female and 20% male), and control groups (78.9% female and 21.1% male). The mean ages of cases and controls were 33.3 ± 4.7 and 35.7 ± 5.1 years, respectively. The gene expression of CB2 in periodontitis was 27.62 ± 7.96 and in healthy subjects was 78.15 ± 23.07. The CB2 was significantly lower than the control group (p = .008). In comparison, the gene expression index of CB1 in the periodontal group (9.42 ± 3.03) was higher than the control group (6.62 ± 1.13) but did not meet a significant value (p = .671). Conclusion The lower expression of CB2 receptors in the periodontitis group may be due to the reduced protective effect of anti‐inflammatory agents. These elements include cannabinoids and the imbalance leading to the predominance of pro‐inflammatory effects. Therefore, the local effects of cannabinoids as an immunomodulator could be useful for oral inflammatory diseases such as periodontitis.

ABSTRACTBackground Preclinical and clinical evidence suggests that cannabis has potential analgesic properties. However, cannabinoid receptor expression and localization within spinal cord pain processing circuits remain to be characterized across sex and species.Aims We aimed to investigate the differential expression of the cannabinoid type 1 (CB1) receptor across dorsal horn laminae and cell populations in male and female adult rats and humans.Methods To investigate and quantify CB1 receptor expression in the spinal dorsal horn across species, we refined immunohistochemical procedures for successful rat and human fixed tissue staining and confocal imaging. Immunohistochemical results were complemented with analysis of CB1 gene (CNR1) expression within rodent and human dorsal horn using single-cell/nuclei RNA sequencing data sets.Results We found that CB1 was preferentially localized to the neuropil within the superficial dorsal horn of both rats and humans, with CB1 somatic staining across dorsal horn laminae. CB1 receptor immunoreactivity was significantly higher in the superficial dorsal horn compared to the deeper dorsal horn laminae for both rats and humans, which was conserved across sex. Interestingly, we found that CB1 immunoreactivity was not primarily localized to peptidergic afferents in rats and humans and that CNR1 (CB1) but not CNR2 (CB2) was robustly expressed in dorsal horn neuron subpopulations of both rodents and humans.Conclusions The conserved preferential expression of CB1 receptors in the superficial dorsal horn in male and female rodents and humans has significant implications for understanding the roles of this cannabinoid receptor in spinal mechanisms of nociception and analgesia.

Abstract Zebrafish is considered an unprecedented animal model in drug discovery. A review of the literature presents highlights and elucidates the biological effects of chemical components found in Cannabis sativa. Particular attention is paid to endocannabinoid system (eCB) and its main receptors (CB1 and CB2). The zebrafish model is a promising one for the study of cannabinoids because of the many similarities to the human system. Despite the recent advances on the eCB system, there is still the need to elucidate some of the interactions and, thus, the zebrafish model can be used for that purpose as it respects the 3Rs concept and reduced time and costs. In view of the relevance of cannabinoids in the treatment and prevention of diseases, as well as the importance of the zebrafish animal model in elucidating the biological effects of new drugs, the aim of this study was to bring to light information on the use of the zebrafish animal model in testing C. sativa-based medicines.

Cannabinoid (CB) receptors (CB1 and CB2) are expressed on cancer cells and their expression influences carcinogenesis in various tumor entities. Cells of the tumor microenvironment (TME) also express CB receptors, however, their role in tumor development is still unclear. We, therefore, investigated the role of TME-derived CB1 and CB2 receptors in a model of non-small cell lung cancer (NSCLC). Leukocytes in the TME of mouse and human NSCLC express CB receptors, with CB2 showing higher expression than CB1. In the tumor model, using CB1- (CB1-/-) and CB2-knockout (CB2-/-) mice, only deficiency of CB2, but not of CB1, resulted in reduction of tumor burden vs. wild type (WT) littermates. This was accompanied by increased accumulation and tumoricidal activity of CD8+ T and natural killer cells, as well as increased expression of programmed death-1 (PD-1) and its ligand on lymphoid and myeloid cells, respectively. CB2-/- mice responded significantly better to anti-PD-1 therapy than WT mice. The treatment further increased infiltration of cytotoxic lymphocytes into the TME of CB2-/- mice. Our findings demonstrate that TME-derived CB2 dictates the immune cell recruitment into tumors and the responsiveness to anti-PD-1 therapy in a model of NSCLC. CB2 could serve as an adjuvant target for immunotherapy.

Abstract Tauopathies are a group of neurodegenerative diseases characterized by the alteration/aggregation of TAU protein, for which there is still no effective treatment. Therefore, new pharmacological targets are being sought, such as elements of the endocannabinoid system (ECS). We analysed the occurrence of changes in the ECS in tauopathies and their implication in the pathogenesis. By integrating gene expression analysis, immunofluorescence, genetic and adeno-associated virus expressing TAU mouse models, we found a TAU-dependent increase in CB2 receptor expression in hippocampal neurons, that occurs as an early event in the pathology and was maintained until late stages. These changes were accompanied by alterations in the endocannabinoid metabolism. Remarkably, CB2 ablation in mice protects from neurodegeneration induced by hTAUP301L overexpression, corroborated at the level of cognitive behaviour, synaptic plasticity, and aggregates of insoluble TAU. At the level of neuroinflammation, the absence of CB2 did not produce significant changes in concordance with a possible neuronal location rather than its classic glial expression in these models. These findings were corroborated in post-mortem samples of patients with Alzheimer’s disease, the most common tauopathy. Our results show that neurons with accumulated TAU induce the expression of the CB2 receptor, which enhances neurodegeneration. These results are important for our understanding of disease mechanisms, providing a novel therapeutic strategy to be investigated in tauopathies.

Background: The endocannabinoid system (ECS) and associated lipid transmitter-based signaling systems play an important role in modulating brain neuroinflammation. ECS is affected in neurodegenerative disorders, such as Alzheimer’s disease (AD). Here we have evaluated the non-psychotropic endocannabinoid receptor type 2 (CB2) and lysophosphatidylinositol G-protein-coupled receptor 55 (GPR55) localization and expression during Aβ-pathology progression. Methods: Hippocampal gene expression of CB2 and GPR55 was explored by qPCR analysis, and brain distribution was evaluated by immunofluorescence in the wild type (WT) and APP knock-in AppNL-G-F AD mouse model. Furthermore, the effects of Aβ42 on CB2 and GPR55 expression were assessed in primary cell cultures. Results: CB2 and GPR55 mRNA levels were significantly upregulated in AppNL-G-F mice at 6 and 12 months of age, compared to WT. CB2 was highly expressed in the microglia and astrocytes surrounding the Aβ plaques. Differently, GPR55 staining was mainly detected in neurons and microglia but not in astrocytes. In vitro, Aβ42 treatment enhanced CB2 receptor expression mainly in astrocytes and microglia cells, whereas GPR55 expression was enhanced primarily in neurons. Conclusions: These data show that Aβ pathology progression, particularly Aβ42, plays a crucial role in increasing the expression of CB2 and GPR55 receptors, supporting CB2 and GPR55 implications in AD.

Childhood absence epilepsy (CAE) is characterized by absence seizures, which are episodes of lack of consciousness accompanied by electrographic spike-wave discharges. About 60% of children and adolescents with absence seizures are affected by major neuropsychological comorbidities, including anxiety. Endocannabinoids and monoamines are likely involved in the pathophysiology of these CAE psychiatric comorbidities. Here, we show that the synthetic cannabinoid receptor type 1/2 (CB1/2R) agonist WIN 55,212-2 (2 mg/kg) has a strain-dependent effect on anxiety-like and motor behavior when assess in the hole board test and cerebral monoaminergic levels in Genetic Absence Epilepsy Rats from Strasbourg (GAERS) and their non-epileptic control (NEC) rat strain. Using quantitative and Temporal pattern (T-pattern) analyses, we found that WIN 55,212-2 did not affect the emotional status of GAERS, but it was anxiolytic in NEC. Conversely, WIN 55,212-2 had a sedative effect in GAERS but was ineffective in NEC. Moreover, vehicle-treated GAERS more motivated to explore by implementing more complex and articulated strategies. These behavioral changes correlate with the reduction of 5-HT in the hippocampus and substantia nigra (SN) and noradrenaline (NA) in the entopeduncular nucleus (EPN) in vehicle-treated GAERS compared to NEC rats, which could contribute to their low anxiety status and hypermotility, respectively. On the other hand, the increased level of NA in the EPN and 5-HT in the SN is consistent with an activation of the basal ganglia output-mediated motor suppression observed in WIN 55,212-2-treated GAERS rats. These data support the view of a strain-dependent alteration of the endocannabinoid system in absence epilepsy by adding evidence of a lower emotional responsiveness and a basal ganglia hypersensitivity to cannabinoids in GAERS compared to NEC rats.

The endocannabinoid system consists mainly of 2-arachidonoylglycerol and anandamide, as well as cannabinoid receptor type 1 and type 2 (CB2). Based on previous studies, we hypothesized that a circulating peptide previously identified as osteogenic growth peptide (OGP) maintains a bone-protective CB2 tone. We tested OGP activity in mouse models and cells, and in human osteoblasts. We show that the OGP effects on osteoblast proliferation, osteoclastogenesis, and macrophage inflammation in vitro, as well as rescue of ovariectomy-induced bone loss and prevention of ear edema in vivo are all abrogated by genetic or pharmacological ablation of CB2. We also demonstrate that OGP binds at CB2 and may act as both an agonist and positive allosteric modulator in the presence of other lipophilic agonists. In premenopausal women, OGP circulating levels significantly decline with age. In adult mice, exogenous administration of OGP completely prevented age-related bone loss. Our findings suggest that OGP attenuates age-related bone loss by maintaining a skeletal CB2 tone. Importantly, they also indicate the occurrence of an endogenous peptide that signals via CB2 receptor in health and disease.

Objective: To investigate the antinociceptive effect of tingenone on inflammatory pain, as well as and the involvement of the cannabinoid receptors type 2 (CB2) and spinal microglia in this process. Methods: Male Swiss mice were subjected to inflammatory pain induced by intraplantar injection of carrageenan. The nociceptive threshold was measured by von Frey filaments test. Tingenone was administered orally 60 min before carrageenan injection. To evaluate the involvement of CB2 receptor, endocannabinoids, and microglia, AM630 (a CB2 receptor antagonist), MAFP (an inhibitor of an enzyme that hydrolyses endocannabinoids), and minocycline (a microglial inhibitor) were given intrathecally 20 min before tingenone administration. In addition, an immunofluorescence assay was used to evaluate CB2 receptor and CD11B (a microglial marker) expression in the spinal cord dorsal horn. Results: Tingenone significantly reduced carrageenan-induced hyperalgesia, which was reversed by pretreatment with AM630. MAFP and minocycline potentiated and prolonged the tingenone- induced antinociception. CD11B expression was increased in the spinal cord dorsal horn of mice with inflammatory pain pretreated with tingenone, which was reduced by AM630, MAFP, and minocycline. Conclusions: CB2 receptors and endocannabinoids participate in the tingenone-induced antinociception which may involve the inhibition of microglia at spinal level.

Sudhir M Baswan,1,* Allison E Klosner,2,* Kelly Glynn,1 Arun Rajgopal,1 Kausar Malik,1 Sunghan Yim,1 Nathan Stern1 1Innovation and Science, Amway Corporation, Ada, MI, 49355, USA; 2Innovation and Science, Nutrilite Health Institute, Amway Corporation, Buena Park, CA, 90621, USA*These authors contributed equally to this workCorrespondence: Sudhir M Baswan Amway Corporation, 7575 Fulton St E, Ada, MI 49355, USATel +1-616-787-0216Email sudhir.baswan@amway.comAbstract: Though there is limited research confirming the purported topical benefits of cannabinoids, it is certain that cutaneous biology is modulated by the human endocannabinoid system (ECS). Receptors from the ECS have been identified in the skin and systemic abuse of synthetic cannabinoids, and their analogs, have also been associated with the manifestation of dermatological disorders, indicating the effects of the ECS on cutaneous biology. In particular, cannabidiol (CBD), a non-psychoactive compound from the cannabis plant, has garnered significant attention in recent years for its anecdotal therapeutic potential for various pathologies, including skin and cosmetic disorders. Though a body of preclinical evidence suggests topical application of CBD may be efficacious for some skin disorders, such as eczema, psoriasis, pruritis, and inflammatory conditions, confirmed clinical efficacy and elucidation of underlying molecular mechanisms have yet to be fully identified. This article provides an update on the advances in CBD research to date and the potential areas of future exploration.Keywords: cannabidiol, CBD, cannabinoids, endocannabinoids, endocannabinoid system, skin, CB1, CB2, FAAH, AEA

Cannabis sativa L. contains more than 100 phytocannabinoids that can interact with cannabinoid receptors CB1 and CB2. None of the cannabinoid receptor ligands is entirely CB1- or CB2-specific. The effects of cannabinoids therefore differ not just because of different potency at cannabinoid receptors but also because they can interact with other non-CB1 and non-CB2 targets, such as TRPV1, GPR55, and GPR119. The most studied phytocannabinoid is Δ9-tetrahydrocannabinol (THC). THC is a partial agonist at both cannabinoid receptors, but its psychotomimetic effect is produced primarily via activation of the CB1 receptor, which is strongly expressed in the central nervous system, with the noteworthy exception of the brain stem. Although acute cognitive and other effects of THC are well known, the risk of irreversible neuropsychological effects of THC needs further research to elucidate the association. Unlike THC, phytocannabinoid cannabidiol (CBD) does not appear to have psychotomimetic effects but may interact with some of the effects of THC if taken concomitantly. CBD administered orally has recently undergone well-controlled clinical trials to assess its safety in the treatment of paediatric epilepsy syndromes. Their findings point to increased transaminase levels as a safety issue that calls for postmarketing surveillance for liver toxicity. The aim of this review is to summarise what is known about acute and chronic toxicological effects of both compounds and address the gaps in knowledge about the safety of exogenous cannabinoids that are still open.

Both metabotropic (CBRs) and ionotropic cannabinoid receptors (ICRs) have implications in a range of neurological disorders. The metabotropic canonical CBRs CB1 and CB2 are highly implicated in these pathological events. However, selective targeting at CB2 versus CB1 offers optimized pharmacology due to the absence of psychoactive outcomes. The ICR transient receptor potential vanilloid type 1 (TRPV1) has also been reported to play a role in CNS disorders. Thus, activation of both targets, CB2 and TRPV1, offers a promising polypharmacological strategy for the treatment of neurological events including analgesia and neuroprotection. This brief research report aims to identify chemotypes with a potential dual CB2/TRPV1 profile. For this purpose, we have rationalized key structural features for activation and performed virtual screening at both targets using curated chemical libraries.

Abstract Background The morbidity and mortality of cancer are significantly impacted by the invasive and metastatic potential of particular subgroups of malignant cells within a tumor. The particular pre‐metastatic properties of cancerous cells are thus a critical target for novel therapeutics in the oncology field. Cannabinoid molecules have been investigated in recent years in the context of invasion and metastasis of various malignancies, with varying effects reported in the literature. Recent Findings There was substantial variability in the findings reported by the literature of the effects of cannabinoid molecules on cancer cell invasion and metastasis. These effects varied depending on which ligand and which of the CB1, CB2, or GPR55 receptors were investigated. These findings suggest a role for the phenomenon of biased signaling in explaining the diversity of effects of cannabinoid molecules on cancer cell invasion. Conclusion While substantially more investigation is required into the effects of cannabinoid molecules on cancer cell invasion and metastasis, we describe in this review the significant diversity in the responses of cancer cells to cannabinoid molecules in terms of their invasive and metastatic capacities.

Research on endocannabinoid signaling has greatly advanced our understanding of how the excitability of neural circuits is controlled in health and disease. In general, endocannabinoid signaling at excitatory synapses suppresses excitability by inhibiting glutamate release, while that at inhibitory synapses promotes excitability by inhibiting GABA release, although there are some exceptions in genetically epileptic animal models. In the epileptic brain, the physiological distributions of endocannabinoid signaling molecules are disrupted during epileptogenesis, contributing to the occurrence of spontaneous seizures. However, it is still unknown how endocannabinoid signaling changes during seizures and how the redistribution of endocannabinoid signaling molecules proceeds during epileptogenesis. Recent development of cannabinoid sensors has enabled us to investigate endocannabinoid signaling in much greater spatial and temporal details than before. Application of cannabinoid sensors to epilepsy research has elucidated activity-dependent changes in endocannabinoid signaling during seizures. Furthermore, recent endocannabinoid research has paved the way for the clinical use of cannabidiol for the treatment of refractory epilepsy, such as Dravet syndrome, Lennox-Gastaut syndrome and tuberous sclerosis complex. Cannabidiol significantly reduces seizures and is considered to have comparable tolerability to conventional antiepileptic drugs. In this article, we introduce recent advances in research on the roles of endocannabinoid signaling in epileptic seizures and discuss future directions.

Abstract Background CB2 (cannabinoid receptor 2) agonists have been shown to exert anti-tumor activities in different tumor types. However, there is no study exploring the role of MDA19 (a novel CB2 agonist) in tumors. In this study we aimed to investigate the effects of MDA19 treatment on HCC cell lines, Hep3B and HepG2 and determine the relevant mechanisms. Results Cell proliferation analysis, including CCK8 and colony formation assays, indicated that MDA19 treatment inhibited HCC cell proliferation in a dose- and time-dependent manner. Flow cytometry suggested that MDA19 induced cell apoptosis and activation of mitochondrial apoptosis pathway. Transwell assay indicated that HCC cell migration and invasion were significantly inhibited by MDA19 treatment. Mechanism investigation suggested that MDA19 induced inactivation of AKT signaling pathway in HCC cells. In addition, we investigated the function of CB2receptor in HCC and its role in the anti-tumor activity of MDA19. By searching on Kaplan-Meier plotter (http://kmplot.com/analysis/), we found that HCC patients with high CB2 expression had a better survival and CB2 expression was significantly associated with gender, clinical stages and race of HCC patients (P

Selective modulation of peripheral cannabinoid receptors (CBRs) has potential therapeutic applications in medical conditions, including obesity, diabetes, liver diseases, GI disorders and pain. While there have been considerable efforts to produce selective antagonists or full agonists of CBRs, there has been limited reports on the development of partial agonists. Partial agonists targeting peripheral CBRs may have desirable pharmacological profiles while not producing centrally mediated dissociative effects. Bayer reported that BAY 59-3074 is a CNS penetrant partial agonist of both CB1 and CB2 receptors with efficacy in rat models of neuropathic and inflammatory pain. In this report, we demonstrate our efforts to synthesize analogs that would favor peripheral selectivity, while maintaining partial agonism of CB1. Our efforts led to the identification of a novel compound, which is a partial agonist of the human CB1 (hCB1) receptor with vastly diminished brain exposure compared to BAY 59-3074.

With implantation, mouse stromal cells begin to transform into epithelial-like cells surrounding the implantation chamber forming an avascular zone called the primary decidual zone (PDZ). In the mouse, the PDZ forms a transient, size-dependent permeable barrier to protect the embryo from maternal circulating harmful agents. The process of decidualization is critical for pregnancy maintenance in mice and humans. Mice deficient in cannabinoid receptors, CB1 and CB2, show compromised PDZ with dysregulated angiogenic factors, resulting in the retention of blood vessels and macrophages. This phenotype is replicated in Cnr1-/- but not in Cnr2-/-mice. In vitro decidualization models suggest that Cnr1 levels substantially increase in mouse and human decidualizing stromal cells, and that neutralization of CB1 signaling suppresses decidualization and misregulates angiogenic factors. Taken together, we propose that implantation quality depends on appropriate angiogenic events driven by the integration of CB2 in endothelial cells and CB1 in decidual cells.

L-dopa induced dyskinesia (LID) is a debilitating side-effect of the primary treatment used in Parkinson's disease (PD), l-dopa. Here we investigate the effect of HU-308, a cannabinoid CB2 receptor agonist, on LIDs. Utilizing a mouse model of PD and LIDs, induced by 6-OHDA and subsequent l-dopa treatment, we show that HU-308 reduced LIDs as effectively as amantadine, the current frontline treatment. Furthermore, treatment with HU-308 plus amantadine resulted in a greater anti-dyskinetic effect than maximally achieved with HU-308 alone, potentially suggesting a synergistic effect of these two treatments. Lastly, we demonstrated that treatment with HU-308 and amantadine either alone, or in combination, decreased striatal neuroinflammation, a mechanism which has been suggested to contribute to LIDs. Taken together, our results suggest pharmacological treatments with CB2 agonists merit further investigation as therapies for LIDs in PD patients. Furthermore, since CB2 receptors are thought to be primarily expressed on, and signal through, glia, our data provide weight to suggestion that neuroinflammation, or more specifically, altered glial function, plays a role in development of LIDs.

The endocannabinoid system is involved in the nociceptive and anti-inflammatory pathways, and a lowered expression of CB2 receptors has been associated with inflammatory conditions, such as osteoarthritis (OA). This suggests that CB2 modulators could be novel therapeutic tools to treat OA. In the present study, the involvement of Harpagophytum procumbens root extract, a common ingredient of nutraceuticals used to treat joint disorders, in CB2 modulation has been evaluated. Moreover, to clarify the effects of the pure single components, the bioactive constituent, harpagoside, and the main volatile compounds were studied alone or in a reconstituted mixture. Human fibroblast-like synoviocytes, extracted by joints of patients, who underwent a total knee replacement, were treated with an H. procumbens root extract dissolved in DMSO (HPEDMSO). The effectiveness of HPEDMSO to affect CB2 pathways was studied by analyzing the modulation of cAMP, the activation of PKA and ERK MAP kinase, and the modulation of MMP-13 production. HPEDMSO was able to inhibit the cAMP production and MAP kinase activation and to down-regulate the MMP-13 production. Pure compounds were less effective than the whole phytocomplex, thus suggesting the involvement of synergistic interactions. Present findings encourage further mechanistic studies and support the scientific basis of the use of H. procumbens in joint disorders.

Abstract Background Women live about 4 years longer due to lower prevalence of cardiovascular complication with ageing. However, the mechanisms involved in the preservation of heart functionality in women have not been fully elucidated. The endocannabinoid system fulfils a significant role in the regulation of cardiovascular system functioning. Cannabinoids, acting through specific receptors (CB1 and CB2), influence on blood pressure, heart rate and myocardial contractility. The function of cardiac muscle cells is strictly dependent on calcium ions. Calcium homeostasis in cardiomyocytes is subjected to complex regulation via calcium-binding proteins. Among them, increasing attention has been paid to the recently discovered S100A6 and CacyBP/SIP. In order to better understand sex differences in the regulation of cardiomyocyte function during ageing, we undertook the present research aimed at immunohistochemical identification and comparative evaluation of cannabinoid receptors, S100A6 and CacyBP/SIP, in the myocardium of ageing men and women. Methods The study was conducted on the hearts of 12 men and 10 women (organ donors) without a history of cardiovascular disease. The subjects were divided into two age groups: subjects older than 50 years and subjects under 50 years old. Paraffin heart sections were processed by immunohistochemistry for detection of cannabinoids receptors, S100A6 and CacyBP/SIP. In the heart samples from each study, participant’s expression of genes coding for CB1, CB2, S100A6 and CacyBP/SIP using real-time PCR method was measured. Results CB1 and CB2 immunoreactivity in the cytoplasm of cardiomyocytes in the heart of subjects over 50 was weaker than in younger individuals. In the heart of younger men, CB1-immunoreactivity was weaker and CB2-immunoreaction was stronger compared to women. In the hearts of older men, the CB1-immunostaining was more intense and CB2-immunoreactivity was weaker than in women. Immunodetection of CB1 shoved the presence of receptor in the intercalated discs, but only in the hearts of individuals over the 50 years old. In the hearts of older individuals, stronger immunolabelling was observed for S100A6 and CacyBP/SIP. Male hearts had greater S100A6-immunoreactivity (both age groups) but less CacyBP/SIP immunostaining (individuals over 50 years) compared to the age-matched women. The expression of genes coding CB1, CB2, S100A6 and CacyBP/SIP in the human heart was sex and age-dependent. Observed changes between men and women as well as between subject under and over 50 years were consistent with immunohistochemically stated changes in peptide content. Conclusion Together, the data presented here indicate a close interaction between ageing and sex on the distribution and levels of cannabinoid receptors (CB1, CB2), S100A6 and CacyBP/SIP in the human heart.

Abstract Background Recent studies have shown a significant role of the endocannabinoid system, apelin and S100A6 protein in the regulation of cardiovascular system functioning. The aim of the study was to compare and evaluate the distribution of cannabinoid receptors (CB1 and CB2), apelin and S100A6 protein in the heart of healthy women in different age groups. Methods The study was conducted on the hearts of 10 women (organ donors) without a history of cardiovascular disease, who were divided into two age groups: women older than 50 years and women under 50 years of age. Paraffin heart sections were processed by immunohistochemistry for detection of cannabinoids receptors (CB1 and CB2), apelin and S100A6 protein. Results CB1 and CB2 immunoreactivity in the cytoplasm of cardiomyocytes in the heart of women over 50 was weaker than in younger individuals. There was also strong immunoreactivity of CB1 in intercalated discs (ICDs) of the heart, only in women over 50. The presence of this receptor in this location was not found in women under 50. Apelin- and S100A6-immunoreactivity in the cardiomyocytes was stronger in older women compared to women under 50.The CB1, apelin and S100A6 immunostaining in the endothelium of myocardial vessels was weaker in women over 50 than in younger women, while intensity of CB2- immunoreaction in coronary endothelium was similar in both groups of women. The results of the study indicate the important role of endocannabinoids, apelin, and S100A6 protein in cardiac muscle function. Conclusion This report might contribute to a better understanding of the role of endocannabinoid system, apelin and S100 proteins in heart function as well as shed new light on processes involved in age-related cardiomyopathy.

Multiple sclerosis (MS) is a chronic and disabling disorder of the central nervous system (CNS) characterized by neuroinflammation leading to demyelination. Recently a combination of Δ9-tetrahydrocannabinol (THC) and Cannabidiol (CBD) extracted from Cannabis has been approved in many parts of the world to treat MS-related spasticity. THC+CBD combination was also shown to suppresses neuroinflammation, although the mechanisms remain to be further elucidated. In the current study, we demonstrate that THC+CBD combination therapy (10 mg/kg each) but not THC or CBD alone, attenuates murine experimental autoimmune encephalomyelitis (EAE) by reducing neuroinflammation and suppression of Th17 and Th1 cells. These effects were mediated through CB1 and CB2 receptors inasmuch as, THC+CBD failed to ameliorate EAE in mice deficient in CB1 and CB2. THC+CBD treatment also caused a decrease in the levels of brain infiltrating CD4+ T cells and pro-inflammatory molecules (IL-17, INF-γ, TNF-α, IL-1β, IL-6, and TBX21), while increasing anti-inflammatory phenotype such as FoxP3, STAT5b, IL-4, IL-10, and TGF-β. Also, the brain-derived cells showed increased apoptosis along with decreased percentage in G0/G1 phase with increased percentage in G2/M phase of cell cycle. miRNA microarray analysis of brain-derived CD4+ T cells revealed that THC+CBD treatment significantly down-regulated miR-21a-5p, miR-31-5p, miR-122-5p, miR-146a-5p, miR-150-5p, miR-155-5p, and miR-27b-5p while upregulating miR-706-5p and miR-7116. Pathway analysis showed that majority of the down-regulated miRs targeted molecules involved in cycle arrest and apoptosis such as CDKN2A, BCL2L11, and CCNG1, as well as anti-inflammatory molecules such as SOCS1 and FoxP3. Additionally, transfection studies involving miR-21 and use of Mir21−/− mice suggested that while this miR plays a critical role in EAE, additional miRs may also be involved in THC+CBD-mediated attenuation of EAE. Collectively, this study suggests that combination of THC+CBD suppresses neuroinflammation and attenuates clinical EAE development and that this effect is associated with changes in miRNA profile in brain-infiltrating cells.

Alzheimer’s disease (AD) is a detrimental brain disorder characterized by a gradual cognitive decline and neuronal deterioration. To date, the treatments available are effective only in the early stage of the disease. The AD etiology has not been completely revealed, and investigating new pathological mechanisms is essential for developing effective and safe drugs. The recreational and pharmacological properties of marijuana are known for centuries, but only recently the scientific community started to investigate the potential use of cannabinoids in AD therapy—sometimes with contradictory outcomes. Since the endocannabinoid system (ECS) is highly expressed in the hippocampus and cortex, cannabis use/abuse has often been associated with memory and learning dysfunction in vulnerable individuals. However, the latest findings in AD rodent models have shown promising effects of cannabinoids in reducing amyloid plaque deposition and stimulating hippocampal neurogenesis. Beneficial effects on several dementia-related symptoms have also been reported in clinical trials after cannabinoid treatments. Accordingly, future studies should address identifying the correct therapeutic dosage and timing of treatment from the perspective of using cannabinoids in AD therapy. The present paper aims to summarize the potential and limitations of cannabinoids as therapeutics for AD, focusing on recent pre-clinical and clinical evidence.

The persistence of human immunodeficiency virus-1 (HIV)-associated neurocognitive disorders (HAND) in the era of effective antiretroviral therapy suggests that modern HIV neuropathogenesis is driven, at least in part, by mechanisms distinct from the viral life cycle. Identifying more subtle mechanisms is complicated by frequent comorbidities in HIV+ populations. One of the common confounds is substance abuse, with cannabis being the most frequently used psychoactive substance among people living with HIV. The psychoactive effects of cannabis use can themselves mimic, and perhaps magnify, the cognitive deficits observed in HAND; however, the neuromodulatory and anti-inflammatory properties of cannabinoids may counter HIV-induced excitotoxicity and neuroinflammation. Here, we review our understanding of the cross talk between HIV and cannabinoids in the central nervous system by exploring both clinical observations and evidence from preclinical in vivo and in vitro models. Additionally, we comment on recent advances in human, multi-cell in vitro systems that allow for more translatable, mechanistic studies of the relationship between cannabinoid pharmacology and this uniquely human virus.

The presented work is aimed at developing a mould geometry suitable for casting both low- and high-alloy steel grades into 500 kg experimental ingots. The high Height-to-Diameter (H/D)-ratio mould currently used in COMTES FHT Inc. served as a reference and for finite element method simulations (FEM) of the filling and solidification process. The optimized mould geometry, balancing the porosity and segregations, was determined using MAGMA software. Four different steel grades were defined for the simulation. Case studies were carried out for 34CrNiMo6 (W.Nr. 1.6582), DHQ8, CB2 and borated stainless steel grades ranging from low-alloy steel to high-alloy steel. Extended user-defined criteria and verified boundary conditions were used to predict the formation of A-segregations in cast steel. Both primary (PDAS) and secondary (SDAS) arm spacings were modelled as well. The optimized mould shape and the casting assembly were designed based on the simulation results.

The endocannabinoid system (ECS) is involved in the modulation of several basic biological processes, having widespread roles in neurodevelopment, neuromodulation, immune response, energy homeostasis and reproduction. In the adult central nervous system (CNS) the ECS mainly modulates neurotransmitter release, however, a substantial body of evidence has revealed a central role in regulating neurogenesis in developing and adult CNS, also under pathological conditions. Due to the complexity of investigating ECS functions in neural progenitors in vivo, we tested the suitability of the ST14A striatal neural progenitor cell line as a simplified in vitro model to dissect the role and the mechanisms of ECS-regulated neurogenesis, as well as to perform ECS-targeted pharmacological approaches. We report that ST14A cells express various ECS components, supporting the presence of an active ECS. While CB1 and CB2 receptor blockade did not affect ST14A cell number, exogenous administration of the endocannabinoid 2-AG and the synthetic CB2 agonist JWH133 increased ST14A cell proliferation. Phospholipase C (PLC), but not PI3K pharmacological blockade negatively modulated CB2-induced ST14A cell proliferation, suggesting that a PLC pathway is involved in the steps downstream to CB2 activation. On the basis of our results, we propose ST14A neural progenitor cells as a useful in vitro model for studying ECS modulation of neurogenesis, also in prospective in vivo pharmacological studies.

Endocannabinoids are lipid mediators, enzymatically synthesized and hydrolyzed, that bind cannabinoid receptors. Together with their receptors and metabolic enzymes, they form the “endocannabinoid system” (ECS). Anandamide (AEA) and 2-arachidonoylglycerol (2-AG) are the main endocannabinoids studied in testis. In this study, using the first wave of spermatogenesis as an in vivo model to verify the progressive appearance of germ cells in seminiferous tubules [i.e., spermatogonia, spermatocytes, and spermatids], we analyzed the expression of the main enzymes and receptors of ECS in rat testis. In particular, the expression profile of the main enzymes metabolizing AEA and 2-AG as well as the expression of cannabinoid receptors, such as CB1 and CB2, and specific markers of mitotic, meiotic, and post-meiotic germ cell appearance or activities have been analyzed by RT-PCR and appropriately correlated. Our aim was to envisage a relationship between expression of ECS components and temporal profile of germ cell appearance or activity as well as among ECS components. Results show that expression of ECS components is related to germ cell progression. In particular, CB2 and 2-AG appear to be related to mitotic/meiotic stages, while CB1 and AEA appear to be related to spermatogonia stem cells activity and spermatids appearance, respectively. Our data also suggest that a functional interaction among ECS components occurs in the testis. Indeed, in vitro-incubated testis show that AEA-CB2 activity affects negatively monoacylglycerol-lipase levels via upregulation of CB1 suggesting a CB1/CB2-mediated relationship between AEA and 2-AG. Finally, we provide the first evidence that CB1 is present in fetal gonocytes, during mitotic arrest.

Exposure to stressful situations is one of the risk factors for the precipitation of several psychiatric disorders, including Major Depressive Disorder, Posttraumatic Stress Disorder and Schizophrenia. The hippocampal formation is a forebrain structure highly associated with emotional, learning and memory processes; being particularly vulnerable to stress. Exposure to stressful stimuli leads to neuroplastic changes and imbalance between inhibitory/excitatory networks. These changes have been associated with an impaired hippocampal function. Endocannabinoids (eCB) are one of the main systems controlling both excitatory and inhibitory neurotransmission, as well as neuroplasticity within the hippocampus. Cannabinoids receptors are highly expressed in the hippocampus, and several lines of evidence suggest that facilitation of cannabinoid signaling within this brain region prevents stress-induced behavioral changes. Also, chronic stress modulates hippocampal CB1 receptors expression and endocannabinoid levels. Moreover, cannabinoids participate in mechanisms related to synaptic plasticity and adult neurogenesis. Here, we discussed the main findings supporting the involvement of hippocampal cannabinoid neurotransmission in stress-induced behavioral and neuroplastic changes.

Hepatic fibrosis is the consequence of an unresolved wound healing process in response to chronic liver injury and involves multiple cell types and molecular mechanisms. The hepatic endocannabinoid and apelin systems are two signalling pathways with a substantial role in the liver fibrosis pathophysiology—both are upregulated in patients with advanced liver disease. Endogenous cannabinoids are lipid-signalling molecules derived from arachidonic acid involved in the pathogenesis of cardiovascular dysfunction, portal hypertension, liver fibrosis, and other processes associated with hepatic disease through their interactions with the CB1 and CB2 receptors. Apelin is a peptide that participates in cardiovascular and renal functions, inflammation, angiogenesis, and hepatic fibrosis through its interaction with the APJ receptor. The endocannabinoid and apelin systems are two of the multiple cell-signalling pathways involved in the transformation of quiescent hepatic stellate cells into myofibroblast like cells, the main matrix-producing cells in liver fibrosis. The mechanisms underlying the control of hepatic stellate cell activity are coincident despite the marked dissimilarities between the endocannabinoid and apelin signalling pathways. This review discusses the current understanding of the molecular and cellular mechanisms by which the hepatic endocannabinoid and apelin systems play a significant role in the pathophysiology of liver fibrosis.

Emerging research demonstrates that exercise, including both acute and chronic exercise, may influence episodic memory function. To date, mechanistic explanations of this effect are often attributed to alterations in long-term potentiation, neurotrophic production, angiogenesis, and neurogenesis. Herein, we discuss a complementary mechanistic model, suggesting that the endocannabinoid system may, in part, influence the effects of exercise on memory function. We discuss the role of the endocannabinoid system on memory function as well as the effects of exercise on endocannabinoid alterations. This is an exciting line of inquiry that should help delineate new insights into the mechanistic role of exercise on memory function.

The aim of review. To analyze main epidemiologic tendencies of development of alcoholic liver disease, mechanisms of its development, mortality in this population of patients. To discuss an option of new targets for treatment of this disease.Original positions. Alcohol abuse is the third leading cause of morbidity and mortality in the young age. In the studies it was demonstrated, that there is significant difference in death rate of liver cirrhosis between European countries. Disease risk factors include: amount of taken alcohol, its type, frequency of use, female gender, excessive body weight. Besides damaging effect, in response to admission of alcohol processes of protection and neogenesis of the liver through activation of IL-6, IL-22 receptors, STAT-proteins system are initiated. Mechanism of TNFα action, resulting in apoptosis of hepatocytes through activation of acidic sphingomyelinase is investigated.Conclusion. Application of acidic sphingomyelinase, influence on Kupffer's cells phenotype, cannabinoids receptors as new targets for treatment of alcoholic liver disease is possible. At treatment of patients with severe alcohol-induced hepatitis corticosteroids considerably increasing 28-day's survival rate are actively applied. To increase efficacy of treatment of acute alcoholic hepatitis the issue of addition to standard treatment by glucocorticosteroids IL-22 is taken into account.

The endocannabinoid system (ECS) has been implicated in many physiological functions, including the regulation of appetite, food intake and energy balance, a crucial involvement in brain reward systems and a role in psychophysiological homeostasis (anxiety and stress responses). We first introduce this important regulatory system and chronicle what is known concerning the signal transduction pathways activated upon the binding of endogenous cannabinoid ligands to the Gi/0-coupled CB1 cannabinoid receptor, as well as its interactions with other hormones and neuromodulators which can modify endocannabinoid signaling in the brain. Anorexia nervosa (AN) and bulimia nervosa (BN) are severe and disabling psychiatric disorders, characterized by profound eating and weight alterations and body image disturbances. Since endocannabinoids modulate eating behavior, it is plausible that endocannabinoid genes may contribute to the biological vulnerability to these diseases. We present and discuss data suggesting an impaired endocannabinoid signaling in these eating disorders, including association of endocannabinoid components gene polymorphisms and altered CB1-receptor expression in AN and BN. Then we discuss recent findings that may provide new avenues for the identification of therapeutic strategies based on the endocannabinod system. In relation with its implications as a reward-related system, the endocannabinoid system is not only a target for cannabis but it also shows interactions with other drugs of abuse. On the other hand, there may be also a possibility to point to the ECS as a potential target for treatment of drug-abuse and addiction. Within this framework we will focus on enzymatic machinery involved in endocannabinoid inactivation (notably fatty acid amide hydrolase or FAAH) as a particularly interesting potential target. Since a deregulated endocannabinoid system may be also related to depression, anxiety and pain symptomatology accompanying drug-withdrawal states, this is an area of relevance to also explore adjuvant treatments for improving these adverse emotional reactions.

Although the mortality rates of cirrhosis are underestimated, its socioeconomic burden has demonstrated a significant global impact. Cirrhosis is defined by the disruption of normal liver architecture after years of chronic insult by different etiologies. Treatment modalities are recommended primarily in decompensated cirrhosis and specifically tailored to the different manifestations of hepatic decompensation. Antifibrogenic therapies are within an active area of investigation. The endocannabinoid system has been shown to play a role in liver disease, and cirrhosis specifically, with intriguing possible therapeutic benefits. The endocannabinoid system comprises cannabinoid receptors 1 (CB1) and cannabinoid receptor 2 (CB2) and their ligands, endocannabinoids and exocannabinoids. CB1 activation enhances fibrogenesis, whereas CB2 activation counteracts progression to fibrosis. Conversely, deletion of CB1 is associated with an improvement of hepatic fibrosis and steatosis, and deletion of CB2 results in increased collagen deposition, steatosis, and enhanced inflammation. CB1 antagonism has also demonstrated vascular effects in patients with cirrhosis, causing an increase in arterial pressure and vascular resistance as well as a decrease in mesenteric blood flow and portal pressure, thereby preventing ascites. In mice with hepatic encephalopathy, CB1 blockade and activation of CB2 demonstrated improved neurologic score and cognitive function. Endocannabinoids, themselves also have mechanistic roles in cirrhosis. Arachidonoyl ethanolamide (AEA) exhibits antifibrogenic properties by inhibition of HSC proliferation and induction of necrotic death. AEA induces mesenteric vasodilation and hypotension via CB1 induction. 2-arachidonoyl glycerol (2-AG) is a fibrogenic mediator independent of CB receptors, but in higher doses induces apoptosis of HSCs, which may actually show antifibrotic properties. 2-AG has also demonstrated growth-inhibitory and cytotoxic effects. The exocannabinoid, THC, suppresses proliferation of hepatic myofibroblasts and stellate cells and induces apoptosis, which may reveal antifibrotic and hepatoprotective mechanisms. Thus, several components of the endocannabinoid system have therapeutic potential in cirrhosis.

A series of pathological events secondary to spinal cord injury (SCI) contribute to the spread of the damage, which aggravates neurological deficits. Here we report that a single dose of the neuroprotective endocannabinoid 2-arachidonoyl glycerol (2-AG) administered early after SCI reduces lesion expansion, which was prevented by simultaneous blockade of both CB1 and CB2 receptors but not by blockade of either receptor alone. Treatment with 2-AG also preserves the white matter around the epicenter of the injury. Moreover, in the preserved white matter, 2-AG protects myelin from damage and reduces oligodendrocyte loss. In addition to these protective actions at the epicenter region, 2-AG also inhibits the myelin damage and delayed oligodendrocyte loss induced at 10 mm from the epicenter. Interestingly, the early protective action of 2-AG is maintained 28 days after injury, when the lesion size is still smaller and the preservation of white matter is better in 2-AG-treated animals. Therefore, our results show that 2-AG protects from the expansion of the lesion and white matter damage, which suggest that this endogenous cannabinoid may be useful as a protective treatment for acute SCI.

The fungal pathogen Candida albicans transforms arachidonic acid (AA) into 3-hydroxyarachidonic acid [3(R)-HETE], and we investigated if its nonpathogenic and 3(R)-HETE-producing close relative, Dipodascopsis uninucleata, could similarly transform the endocannabinoid/endovanilloid anandamide into 3-hydroxyanandamide (3-HAEA). We found that D. uninucleata converts anandamide into 3-HAEA, and we therefore developed an enantiodivergent synthesis for this compound to study its pharmacological activity. Both enantiomers of 3-HAEA were as active as anandamide at elevating intracellular Ca2+ via TRPV1 receptors overexpressed in HEK-293 cells, while a∼70–90-fold and ∼45–60-fold lower affinity at cannabinoid CB1 and CB2 receptors was instead observed. Patch clamp recordings showed that 3(R)-HAEA activates a TRPV1-like current in TRPV1-expressing HEK-293 cells. Thus, 3(R)-HETE-producing yeasts might convert anandamide released by host cells at the site of infection into 3(R)-HAEA, and this event might contribute to the inflammatory and algogenous responses associated to fungal diseases.

Endocannabinoids are lipid mediators with protective effects in many diseases of the nervous system. We have studied the modulation of the endocannabinoid system after a spinal cord contusion in rats. In early stages, lesion induced increases of anandamide and palmitoylethanolamide (PEA) levels, an upregulation of the synthesizing enzyme NAPE-phospholipase D and a downregulation of the degradative enzyme FAAH. In delayed stages, lesion induced increases in 2-arachidonoylglycerol and a strong upregulation of the synthesizing enzyme DAGL-α, that is expressed by neurons, astrocytes and immune infiltrates. The degradative enzyme MAGL was also moderately increased but only 7 days after the lesion. We have studied the cellular targets for the newly formed endocannabinoids using RT-PCR and immunohistochemistry against CB1 and CB2 receptors. We observed that CB1 was constitutively expressed by neurons and oligodendrocytes and induced in reactive astrocytes. CB2 receptor was strongly upregulated after lesion, and mostly expressed by immune infiltrates and astrocytes. The endocannabinoid system may represent an interesting target for new therapeutical approaches to spinal cord injury.

Cnr2 is one of two cannabinoid receptors known to regulate bone metabolism. Here, we compared the whole bone properties of femora and tibiae from three-month-old Cnr2−/- mice with wild-type controls using a C57BL/6 background. Bending stiffness was measured by three-point bending. The elastic modulus, density and mineral content were measured using ultrasound, Archimedes’ principle and ashing. Micro-CT was used to measure the second moment of area, inner and outer perimeters of the cortical shaft and trabecular parameters. Deleting Cnr2 increased the bending stiffness by increasing the second moment of area. Bone from affected male mice had a greater modulus than controls, although no difference was observed in females. The fractional volume of trabecular bone was greater in Cnr2−/- females than controls, while no difference was seen in males. These data indicate that inactivating Cnr2 increases the amount of cortical bone in both males and females at 3 months of age, but the effect on trabecular bone is different in the two sexes. These findings extend previous studies looking only at trabecular bone and provide further support for the possible use of Cnr2 antagonists for improving bone properties that may be of value in the treatment of bone disorders.

Aim: Here, we aimed to explore cannabinoid receptor 2 (CB2) expression in ovine ileum during prenatal development using immunohistochemistry. Materials and Methods: We used 20 ileal samples from fetal sheep whose gestational ages range from 63 to 147 days. To visualize CB2 expression in ileum, we applied labeled Streptavidin Biotin method to paraffin sections. Results: The majority of the intestinal epithelium showed a weak positive reaction for CB2. Interestingly, we observed intense intracytoplasmic staining in some epithelial cells of both intestinal villi and crypts. Moreover, CB2 immunoreaction was observed in ganglion cells in submucosal and myenteric plexus. We also determined positive staining in smooth muscle cells of lamina and tunica muscularis. Towards the end of the prenatal period, we found histologically mature Peyers patches. CB2 staining was detected in the majority of immune cells forming the follicle and dome region. Follicle associated epithelium also showed CB2 immunoreaction. Conclusion: The present study is the first to describe CB2 expression in fetal sheep ileum. CB2 expression was determined in epithelial cells, smooth muscles, enteric neurons and Peyers patches in ileum fetal sheep under physiological conditions. Considering the morphology of epithelial cells with intense CB2 staining and previous findings, CB2 may be a possible marker for enteroendocrine cells during the prenatal period.