Effects Of Cannabinoid Receptor Research Articles

Activation of cannabinoid receptors in breast cancer cells improves osteoblast viability in cancer-bone interaction model while reducing breast cancer cell survival and migration.

The endocannabinoid system has been postulated to help restrict cancer progression and maintain osteoblastic function during bone metastasis. Herein, the effects of cannabinoid receptor (CB) type 1 and 2 activation on breast cancer cell and osteoblast interaction were investigated by using ACEA and GW405833 as CB1 and CB2 agonists, respectively. Our results showed that breast cancer cell (MDA-MB-231)-derived conditioned media markedly decreased osteoblast-like UMR-106 cell viability. In contrast, media from MDA-MB-231 cells pre-treated with GW405833 improved UMR-106 cell viability. MDA-MB-231 cells were apparently more susceptible to both CB agonists than UMR-106 cells. Thereafter, we sought to answer the question as to how CB agonists reduced MDA-MB-231 cell virulence. Present data showed that co-activation of CB1 and CB2 exerted cytotoxic effects on MDA-MB-231 cells by increasing apoptotic cell death through suppression of the NF-κB signaling pathway in an ROS-independent mechanism. ACEA or GW405833 alone or in combination also inhibited MDA-MB-231 cell migration. Thus, it can be concluded that the endocannabinoid system is able to provide protection during breast cancer bone metastasis by interfering cancer and bone cell interaction as well as by the direct suppression of cancer cell growth and migration.

Cannabinoid receptors modulate LPS-induced increase of class-II transactivator expression levels in a microglial cell line

Microglial antigen generation (MAG) is an essential process in regulating disease states and homeostasis of the central nervous system. MAG is considered as responsible autoimmune mechanism in neurodegenerative diseases such as Alzheimer's, Parkinson's, and Huntington's diseases. Neuroprotective and regulator effects of cannabinoid receptors on these disease states and modulation with pharmacological agents are urgent subjects in recent decades. Although different aspects of microglial immune response have been investigated, specific effects of these receptor subtypes in the MAG are still unclear. Therefore, in the current study, we have investigated the effects of CB1 and CB2 receptors on antigen generation by investigating MHC-II and its master regulator CIITA by specific cannabinoid agents (ACEA, AM-251, CP 55,940, and SR144528) in the LPS-induced BV-2 cells. Additionally, the effects of drug treatments on inflammatory status were measured by determining IL-1β, IL-6, and TNF-α levels. LPS-induced increase in MHC-II and CIITA expression was inhibited by specific CB1 agonist, ACEA, and nonselective cannabinoid agonist CP 55,940. A combination with specific CB1 antagonist AM-251 prevented these inhibitory effects of ACEA and CP 55,940 on both MHC-II and CIITA expression. Although specific CB2 antagonist, SR144528, also prevented the inhibitory effect of CP 55,940 on MHC-II, it did not affect CIITA expression. LPS-induced IL-1β, IL-6, and TNF-α increase both attenuated with CP 55,940 and ACEA treatments. Although both selective cannabinoid antagonists inhibited this effect, preventive effects were more dominant on CB1 receptors. Our results demonstrated that CB1 receptors majorly mediates LPS-induced MHC-II and its regulator CIITA expression in microglial cells.

Effects of cannabinoid receptor 2 agonist AM1241 on proliferation, activation and apoptosis ofHSC-T6 cells induced by TGF-β1

Effects of cannabinoid receptor 2 agonist AM1241 on proliferation, activation and apoptosis ofHSC-T6 cells induced by TGF-β1

Effects of cannabinoid receptor 2 synthetic agonist, AM1241, on bleomycin induced pulmonary fibrosis

ABSTRACT Bleomycin (BLM) is a chemotherapeutic agent that can cause pulmonary fibrosis. Little is known about the possible protective role of the CB2 receptor agonist, AM1241. We investigated the effects of CB2 receptor activation by AM1241 on BLM induced lung fibrosis in a rat model. BLM was administered via the trachea. Adult female Wistar rats were divided into five groups: saline (control group), BLM (BLM group), CB2 agonist (AM1241) + BLM (BLMA group), CB2 antagonist (AM630) and CB2 agonist (AM1241) + BLM (BLMA + A group), and vehicle (dimethylsulfoxide) + BLM (BLM + vehicle group). Hydroxyproline, collagen type 1, total protein, glutathione (GSH), malondialdehyde (MDA), interleukin (IL)-6 and tumor necrosis factor (TNF)-α levels were measured in lung fibrosis and control tissue using standard methods. We investigated the histopathology of lung tissue to determine the extent of fibrosis. We found significantly higher levels of hydroxyproline, TNF-α, IL-6 and total protein in the BLM group compared to the BLMA group. The level of GSH also was higher in the BLMA group compared to the BLM group. Inflammation and fibrotic changes were significantly reduced in the BLMA group. Our findings suggest that CB2 receptor activation provided protection against BLM induced pulmonary fibrosis by suppressing oxidative stress and increasing cytokines.

The effect of cannabinoid receptor 1 blockade on hepatic free fatty acid profile in mice with nonalcoholic fatty liver disease

We used rimonabant to investigate the role of CB1 receptor on hepatic FFAs profile during NAFLD. Male mice C57BL/6 were divided into: control group fed with control diet 20 weeks (C; n=6); group fed with HFD 20 weeks (HF; n=6); group fed with control diet and treated with rimonabant after 18 weeks (R; n=9); group fed with HFD and treated with rimonabant after 18 weeks (HFR; n=10). Rimonabant (10mg/kg) was administered daily to HFR and R group by oral gavage. Rimonabant decreased liver palmitic acid proportion in HFR group compared to HF group (p<0.05). Liver stearic and oleic acid proportions were decreased in R group compared to control (p<0.01 respectively). Rimonabant increased liver linoleic and arachidonic acid proportions in HFR group compared to HF group (p<0.01 respectively). CB1 blockade may be useful in the treatment of HFD-induced NAFLD due to modulation of plasma lipid and hepatic FFA profile.

The effect of cannabinoid receptor 2 agonist JWH015 on the hyperalgesia induced by remifentanil

Objective To investigate the effects of cannabinoid receptor 2 (cannabinoid receptor 2, CB2R) agonist JWHOl5 on the hyperalgesia induced by remifentanil in a rat model of postoperative pain. Methods Sixty SD rats were randomly divided into 10 groups (n=6 each): control groups (C1 and C2), incisional pain groups (I1 and I2), incisional pain plus JWHOl5 groups (QI and FI), remifentanil groups (R1 and R2), and JWHOl5 plus remifentanil groups (QR and FR). Rats in group QI/QR and FI/FR were intrathecal injection with 10μg JWHOl5 (diluted in 10μl 20% DMSO solution) and intraperitoneal administration with 100μg JWHOl5 (diluted in 10μl 4% DMSO solution) respectively 30 min before plantar incision while rats in group C, I and R were received with the same volume of DMSO solution. Plantar incision surgery was operated in rats of group I, R, QI/FI, and QR/FR. In group R and QR/FR, remifentanil (0.04 mg/kg) was infused subcutaneously to rats with a pump for 30 min at the moment of surgical incision. The paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency (PWTL) at 24 h before incision and at 2 h, 6 h, 24 h and 48 h after incision were tested to evaluate the behavioral changes. Results Compared with group C and baseline, the level of PWMT and PWTL decreased at 2 h, 6 h, 24 h and 48h after incision in group I (P<0.01); Compared with group I , the significant decrease of PWMT and PWTL were observed after incision in group R (P<0.05); Compared with group R, the significant increase of PWMT (7.78±1.09) and PWTL (17.28±1.58) were observed from 6 h after incision in group QR(P<0.05). And the increase of PWMT (7.79±0.72, 9.50±1.17, 7.86±1.16) and PWTL (16.23±1.50, 19.53±1.63, 18.10±0.93) were observed at 6 h, 24 h and 48 h after incision in group FR(P<0.05). Conclusion Intrathecal and intraperitoneal administration of JWHOl5 in this investigation dose could relief remifentanil-induced hyperalgesia in a rat model of postoperative pain. Key words: Cannabinoid; Cannabinoid receptor 2; JWH015; Remifentanil; Hyperalgesia

O037 : Macrophage autophagy mediated the beneficial effects of cannabinoid receptor 2 in alcoholic liver disease

O037 : Macrophage autophagy mediated the beneficial effects of cannabinoid receptor 2 in alcoholic liver disease

Effects of CB1 receptor blockade on monosodium glutamate induced hypometabolic and hypothalamic obesity in rats

Effects of cannabinoid receptor 1 (CB1R) blockade were observed by comparing 9-day and 6-week SR141716 treatments in monosodium glutamate (MSG)-induced hypometabolic and hypothalamic obesity (HO) in rats for the first time and molecular mechanisms were investigated. Compared with normal rats, the MSG rats display typical symptoms of the metabolic syndrome, i.e., excessive abdominal obesity, hypertriglyceridemia, hyperinsulinemia, insulin resistance, and hepatic steatosis, but with lower food intake. Although both the 9-day and 6-week treatments with the specific CB1R antagonist SR141716 effectively lowered body weight, intraperitoneal adipose tissue mass, serum triglyceride (TG), and insulin level, the effect of chronic treatment is more impressive. Moreover, serum cholesterol, free fatty acids (FFA), fasted and postprandial blood glucose, and insulin insensitivity were more effectively improved by 6-week exposure to SR141716, whereas hypophagia was only effective within the initial 2weeks. In addition, hepatic steatosis as well as hepatic and adipocyte morphology was improved. Western blot analysis revealed that the markedly increased CB1R expression and decreased insulin receptor (INR) expression in liver and adipose tissues were effectively corrected by SR141716. Consistent with this, deregulated gene expression of lipogenesis and lipolysis as well as glucose metabolic key enzymes were also restored by SR141716. In conclusion, based on present data we found that: (1) alteration of the hypothalamus in MSG rats leads to a lower expression of INR in crucially insulin-targeted tissues and hyperinsulinemia that was reversed by SR141716, (2) the abnormally increased expression of CB1R in liver and adipose tissues plays a vital role in the pathophysiological process of MSG rats, and (3) chronic CB1R blockade leads to a sustained improvement of the metabolic dysfunctions of MSG rats.

Inhibitory effects of AM630 on inflammatory osteolysis induced by titanium particles

Objective To explore the effect of cannabinoid receptor 2 (CB2) selective antagonist (AM630) on the regulation of osteoclast differentiation from a murine macrophage cell line (RAW264.7)stimulated with titanium(Ti) particles and the receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL).Methods The effect of AM630 and Ti particles on RAW264.7 cell viability was examined by using the methyl thiazol tetrazolium (MTT) assay.Osteoclast formation was measured by tartrate resistant acid phosphatase (TRAP) staining using a commercial kit.The mRNA levels of CB2,creatine phosphokinase(CPK) and NF-κB receptor activation factor(RANK) were detected by using quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR).Enzyme-linked immunosorbent assay (ELISA)was performed to determine interleukin (IL)-1β and tumor necrosis factor (TNF)-t.Results AM630 (0-200 nmol/L) did not affect viability of RAW264.7 cells cultured with Ti particles.The mature osteoclasts were obtained from RAW264.7 cells stimulated with RANKL and Ti particles for 6 days and detected by TRAP staining.AM630 at a doses of ≥ 100 nmol/L significantly reduced the number of TRAP-positive cells as compared with controls.After culture for 6 days,mRNA levels of CB2,RANK and CPK were significantly higher in the media with RANKL and Ti particles than in those without RANKL and Ti particles.When 100 nmol/L AM630 in combination with RANKL and Ti particles was added to the culture system,mRNA levels of CB2,RANK and CPK were significantly decreased.The concentrations of IL-1β and TNF-α in RAW264.7 cell cultures were measured by using ELISA.After culture for 24 h,IL-13 and TNF-α concentrations were significantly increased in the media with Ti particles as compared with those in the media withou Ti particles.The cytokine concentration was further increased with culture time.Conclusion AM630 could significantly inhibit Ti particle-induced inflammatory osteoclastogenesis. Key words: Osteoclasts ; Inflammation ; Titanium particles ; Cannabinoid receptor 2

Cannabinoid Receptor 2 Protects against Acute Experimental Sepsis in Mice

The systemic inflammatory response syndrome can be self-limited or can progress to severe sepsis and septic shock. Despite significant advances in the understanding of the molecular and cellular mechanisms of septic shock, it is still one of the most frequent and serious problems confronting clinicians in the treatments. And the effects of cannabinoid receptor 2 (CB2R) on the sepsis still remain undefined. The present study was aimed to explore the role and mechanism of CB2R in acute sepsis model of mice. Here, we found that mice were more vulnerable for lipopolysaccharide- (LPS-) induced death and inflammation after CB2R deletion (CB2R−/−). CB2R agonist, GW405833, could significantly extend the survival rate and decrease serum proinflammatory cytokines in LPS-treated mice. GW405833 dose-dependently inhibits proinflammatory cytokines release in splenocytes and peritoneal macrophages as well as splenocytes proliferation, and these effects were partly abolished in CB2R−/− splenocytes but completely abolished in CB2R−/− peritoneal macrophages. Further studies showed that GW405833 inhibits LPS-induced phosphorylation of ERK1/2 and STAT3 and blocks IκBα degradation and NF-κB p65 nuclear translocation in macrophages. All data together showed that CB2R provides a protection and is a potential therapeutic target for the sepsis.

The effect of cannabinoid receptor 2 agonist JWH015 on the hyperalgesia induced by remifentanil

Af filiated Drum-Tower Hospital of Medical College of Nanjing Universit y, Department of Anaesthesiology, Nanjing, China

Effect of intrathecal injection JWH015 on the remifentani-induced hyperalgesia in rats

Objective To investigate the effect of cannabinoid receptor 2 (CB2R) agonist JWH015 on the hyperalgesia induced by remifentanil in a rat model of incision pain. Methods Thirty Sprague-Dawley (SD) rats were randomly divided into 5 groups (n=6):control group(group C),incisional pain group(group I),incisional pain plus JWH015 group (group JI),remifentanil group (group R),and JWH015 plus remifentanil group (group JR).10 μg JWH015 diluted by 10 μl 20％ dimethyl sulfoxide (DMSO)solution was intrathecally injected in each rat of group JI and JR 30 min before plantar incision,while group C,I and R received the equivalent amount of normal saline 20％ DMSO 10 μl in the same way.Plantar incision surgery was operated in group I,R,JI,and JR.In group R and JR,remifentanil (0.04 mg/kg,0.4 ml) was infused subcutaneously with a pump for 30 min at the moment of surgical incision.The other groups were received the equivalent amount of normal saline (0.4 ml) instead of remifentanil.The paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency (PWTL) were measurd one day before and at 2,6,24 and 48 h after incision. Results Compared with group C and the baseline,the level of PWMT and PWTL significantly decreased at 2,6,24 and 48 h after incision in group I (P＜0.05); Compared with group I,the significant decrease of PWMT (6.3±0.8),(6.4±0.8),(6.3± 1.0),(6.8±0.9) g and PWTL( 12.8± 1.2),(12.2±0.9),( 13.4± 1.1 ),( 13.5± 1.3 ) s were observed after incision in group R (P＜0.05).Compared with group R,the significant increase of PWMT (7.9±1.0),(9.9±1.1),(8.4±1.1) g and PWTL (17.3±1.9),(19.9±1.3),(17.7±1.2) s were observed at 6,24 and 48 h after incision in group JR (P＜0.05). Conclusions Intrathecal injection of JWH015 can be effectively alleviated remifentanil-induced hyperalgesia in a rat model of incision pain. Key words: Cannabinoids; Cannabinoid receptor 2; JWH015; Remifentanil; Hyperalgesia

Effects of a Cannabinoid Receptor 2 Selective Antagonist on the Inflammatory Reaction to Titanium Particles In Vivo and In Vitro

Wear particle-induced inflammation is a major factor contributing to aseptic loosening in peri-prosthetic tissue. The effects of cannabinoid receptor 2 (CB(2)) on wear particle-induced inflammation remain unclear. Reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay were used to assess the effects of a CB(2)-selective antagonist, AM630, on regulation of the inflammatory reaction and production of pro-inflammatory cytokines in response to in vitro and in vivo stimulation with titanium particles. In vitro studies, in a model for pre-osteoclast-like cells, demonstrated that AM630 inactivation of CB(2) profoundly inhibited interleukin (IL)-1β and tumour necrosis factor (TNF)-α production by RAW264.7 cells stimulated with titanium particles. In vivo findings in a murine air-pouch model of titanium-induced inflammatory osteolysis indicated that AM630 reduced titanium-induced tissue inflammation, seen as a reduction in pouch membrane thickness, inflammatory infiltration and levels of the pro-inflammatory cytokines IL-1β and TNF-α. Thus, inactivation of CB(2) by AM630 inhibited the titanium particle-induced inflammatory reaction by reducing pro-inflammatory cytokines in vitro and in vivo.

Rapid alteration of stress-induced hypothalamic–pituitary–adrenal hormone secretion in the rat: A comparison of glucocorticoids and cannabinoids

The hypothalamic–pituitary–adrenal (HPA) axis self-regulates through a glucocorticoid negative feedback mechanism that is stereotypically slow and long lasting. Rapid (seconds to minutes) glucocorticoid feedback, however, inhibits stress-induced adrenocorticotropic hormone (ACTH) secretion too quickly to result from classic transcriptional effects of the occupied glucocorticoid receptor. Cannabinoids may act as rapid intermediary messengers between glucocorticoids and HPA activation via retroactive inhibition of afferent glutamate stimulation of the corticotropin-releasing factor neurons in the paraventricular nucleus. We demonstrated fast feedback effects of GR stimulation and blockade and observed the effect of cannabinoid receptor (CB1) antagonist AM251 on HPA axis reactivity in vivo. Rats were injected intraperitoneally with varying doses of the specific GR agonist RU28362, the GR antagonist RU486, or AM251 2 min before restraint. Blood was collected at predetermined times and corticosterone and ACTH concentrations were measured. RU28362 blunted stress-induced ACTH secretion while RU486 and AM251 significantly increased stress-induced ACTH release 15 min after restraint onset. Next, we injected AM251 58 min before RU28362, 2 min before restraint, to determine if inhibition of ACTH by RU28362 was contingent on CB1 activation. Unexpectedly, CB1 blockade failed to prevent glucocorticoid negative feedback and instead enhanced it. These studies not only establish an in vivo fast feedback model but show that rapid glucococorticoid negative feedback is similarly altered by GR and CB1 blockade. Although the hormonal consequences of acute AM251 treatment were strikingly similar to those of RU486 treatment, we are unable to draw conclusions about the serial nature of the interaction between GR activation and CB release from these results.

Effects of cannabinoid receptors on skeletal muscle oxidative pathways

The endocannabinoids, a recently discovered endogenous, lipid derived, signaling system regulating energy metabolism, have effects on central and peripheral energy metabolism predominantly via the cannabinoid receptor type 1 (CB1). CB1 is expressed centrally in the hypothalamus and nucleus accumbens and peripherally in adipocytes and skeletal muscle. This study determined the effect of endocannabinoids on the expression of genes regulating energy metabolism in human skeletal muscle. Primary cultures of myotubes (lean and obese; n = 3/group) were treated with the cannabinoid receptor agonist, anandamide (AEA) (0.2 and 5 μM) and the CB1 specific antagonist AM251 (0.2 and 5 μM) separately and in combination for 24 h. The expression of mRNA for AMP-activated protein kinase (AMPK) alpha 1 (α1) and alpha 2 (α2), pyruvate dehydrogenase kinase 4 (PDK4) and peroxisome proliferator-activated receptor-γ co-activator-1α (PGC-1α) were determined using ‘Real Time’ RT-PCR. AMPKα1 mRNA increased in lean and obese myotubes in response to AM251 ( P < 0.05). AEA inhibited the effect of AM251 on AMPKα1 mRNA levels in myotubes from lean and obese subjects ( P < 0.05); the dose–response curve was shifted to the left in the obese. In response to AM251, irrespective of the presence of AEA, PDK4 expression was decreased in lean and obese myotubes ( P < 0.05). Taken together these data suggest that endocannabinoids regulate pathways affecting skeletal muscle oxidation, effects particularly evident in myotubes from obese individuals.