How cannabinoids influence immune function has been examined extensively in the last 30 years. Studies on drug-abusing humans and animals, as well as in. The cannabinoid system and cytokine network. effects possibly through inhibiting the production and action of TNF-alpha and other acute phase cytokines. Cytokines are the signaling proteins synthesized and anti-inflammatory effects of THC were demonstrated in.
action cytokines Cannabinoid on
Surprisingly, a new mechanism of cell death in cancer cells has been demonstrated, which has been related to the activation of autophagic pathway 13 , Originally known as a housekeeping process responsible for the protein and organelle turnover, autophagy has also been indicated to play a significant role in cell death. Autophagic process is achieved through two pathways: They differ in the transport of the material to be degraded within the lysosomal lumen, the type of transported material and its regulation.
Microautophagy has been traditionally regarded as a form of active autophagy to ensure the turnover of long-lived proteins in basal conditions During microautophagy, lysosomes incorporate and digest regions of cytosol, including proteins and cytoplasmic organelles, by invagination or protrusion without the intermediate formation of autophagic vacuoles.
Macroautophagy or autophagy is responsible for the degradation of soluble proteins and organelles under stress conditions It involves the formation of double-membrane vacuoles autophagosomes that sequester portions of the cytoplasm and translocate them to the lysosomes Some, such as Atg1, are involved in upstream steps of autophagosome formation; others, such as Beclin-1 Atg6 , perform their role as part of a core complex that contains vacuolar sorting protein 34 VPS34 ; others, such as LC-3, a protein which is lipidated and inserted into the outer and inner surface of autophagosomes, are essential for the biogenesis of the large double-membrane vesicles Once digested by autophagosome, the substances are reabsorbed in the vacuole and made available to the cells.
At the molecular level, autophagy is regulated by many protein factors which are involved in key steps of the mechanism. This inhibition allows mTOR to act freely thus inhibiting autophagy The other crucial pathway involved in autophagy is sustained by AMPK. This event is responsible for mTOR inhibition and p53 phosphorylation, inducing increase in autophagy and arrest of cell cycle progression, respectively In an attempt to explain the exact role of autophagy in cancer process, Eisenberg-Lerner and Kimchi 21 recently presented a model in which the role of autophagy is dependent on the stage of cell life, i.
In the first two stages autophagy could accelerate tumor development whereas in solid tumors, when apoptosis is absent, autophagy program could be antitumorigenic. It has been demonstrated that autophagy can undertake a complex interplay with apoptosis In relation to cell type, it could serve as a cell survival pathway suppressing apoptosis, or in others, it could trigger a death pathway either in collaboration with apoptosis or substituting to that when the former is defective.
It is very difficult to distinguish the two pathways because some death stimuli can activate both of them and which pathway will be undertaken depends on the molecular expression profile of the cell type An interesting observation concerns the role explained by ER-stress response in determining cell fate. Different injuries, such as calcium imbalance, oxidative stress or misfolded proteins, trigger a condition indicated as stress of endoplasmic reticulum ER-stress which can lead to different cell responses.
ER-stress can drive mitochondria-dependent apoptosis when there is the activation of the transcription factor CHOP; or it can determine cell survival when it is accompanied by the increase in the level of GRP78, the main intraluminar HSP70 that protects cells from the injury. The cross-talk between apoptosis and autophagy is therefore quite complex, and sometimes contradictory, but surely critical to the overall cell fate.
In the last few years, investigations have suggested the potential application of cannabinoids as antitumor drugs because in many in vivo and in vitro tumor models the activation of the cannabinoid system induces cell cycle arrest, inhibition of cell survival and activation of programmed cell death signalling Apart from regulating tumor cell growth and death, other antitumorigenic mechanisms of cannabinoids are currently emerging as a focus of research work, such as their effects on tumor neovascularization, cell migration, adhesion, invasion and metastasis The present review focuses on the impact of cannabinoids on apoptosis and autophagy, the two main death pathways responsible for cancer cell death.
However, there are still many obscure sides on death pathways activated by these compounds and, in particular, on the different contribution of apoptosis and autophagy in cell death.
The anticancer potential of this class of compounds can be very different in the various tumor systems. This depends on the mechanism used by cannabinoids to interact with the cells, i. Regarding the interaction with the target cell, it has been demonstrated that cannabinoids can interact with the specific type 1 or 2 cannabinoid receptors CB1 and CB2 , which are differently expressed in neural and peripheral tissues, respectively, with transient receptor potential channels of the vanilloid type-1 TRPV1 , or directly with membrane microdomains rich in cholesterol named lipid rafts The interaction of cannabinoids with the different receptor types often leads to the same cell fate even if different intracellular signalling cascades can be activated.
For example, the activation of CB receptors can be responsible for the increase in the level of ceramide or the decrease of cAMP level via inhibition of adenilate cyclase. These events result in canonical apoptosis mediated by the activation of caspase activities 29 , It has also been demonstrated that the interaction of cannabinoids with TRPV receptors causes the activation of overlapping mechanisms, such as the mitochondrial apoptotic pathway, although in this case the event is accompanied by the increase in the level of ROS with the consequent oxidative stress However, sometimes the effects of cannabinoids can not be counteracted by the addition of selective cannabinoid receptor antagonists, hypothesizing that another type of interaction between cannabinoids and cell membrane can be present.
In hepatic stellate cells, an interaction between anandamide and lipid rafts, the membrane micro-domains rich in cholesterol, was found. Differently, in hepatocellular carcinoma cells, as well as in melanoma cells, the interaction of the synthetic cannabinoid WIN55, to lipid rafts is able to promote a canonical caspase-dependent apoptotic pathway 33 , Moreover, it is not possible to exclude that the activation of CB1 receptors and the consequent induction of cannabinoid signalling could be mediated by the association of receptor with lipid rafts, as recently demonstrated by Oddi et al 35 in elegant experiments of site-specific mutations of palmitoylated cysteine residue of CB1 receptors.
An exception to that previously reported is represented by GPR55, a novel potential cannabinoid receptor, whose activation is strictly related to cell proliferation The analysis of intracellular mediators of apoptotic cell death induced by cannabinoids leads to the identification of some molecules, such as ceramide, reactive oxygen species ROS , mitogenic kinases MAPK and some survival factors, that, more than others, seem to be representative of cannabinoid actions.
Ceramide is one of the first molecules identified as a mediator of cannabinoid action. The induction of ceramide accumulation mediated by CB receptors leads to apoptosis in mantle cell lymphoma, glioma, colon and pancreatic cancer 37 — The increase in ceramide level can be dependent on the de novo synthesis or the release from membrane sphyngolipids induced by the activation of sphyngomyelinase.
The different origins of ceramide can be disclosed by the employment of specific inhibitors of the two enzymes, fumonisin B1, the inhibitor of ceramide synthase, or desipramine, an inhibitor of sphyngomyelinase 41 , In many cannabinoid signalling pathways, ROS can exert a crucial role in activating both initiator and executioner caspase activities suggesting that continuous oxidative stress can occur following cannabinoid exposure. The involvement of ROS in cannabinoid-induced apoptosis has been widely evidenced in glioma and leukemia cells 43 , Moreover, the demonstration of ROS involvement in cannabinoid-induced apoptosis is also confirmed by the employement of N-acetyl-cysteine, a thiol antioxidant that scavenges ROS, or the NAD P H oxidase inhibitors, which are able to attenuate cannabinoid effects.
A strong interplay between ceramide and generation of oxidative stress has been demonstrated. Ceramide stimulates the formation of ROS and apoptotic mechanisms, establishing a link between sphingolipid metabolism and oxidative stress. Moreover, inhibition of ROS generating enzymes or treatment with antioxidants impairs sphingomyelinase activation and ceramide production.
Therefore, it is plausible to hypothesize that the contemporaneous activation of CB and TRP vanilloid receptors, responsible for the triggering of the two intracellular mediators ceramide and ROS , can augment the effects induced by cannabinoids carrying out an amplification of caspase cascades.
The study on downstream events following ROS generation or ceramide induction has evidenced the involvement of ER-stress. On the other hand, a relationship between the increase in ceramide level and ER-stress is also well documented in cannabinoid-induced apoptosis in tumor cells A plethora of data reports the activation of stress-activated protein kinases or extracellular-related signal kinases in cannabinoid-dependent control of cancer cell growth and survival 43 , Interestingly, it has been demonstrated that the duration of the stimulus can be fundamental for the type of cellular response.
A transient activation of ERK cascade leads to cell survival and proliferation, while long-term ERK activation results to apoptotic response as proposed by Galve-Roperh et al 48 in malignant glioma and confirmed in Kaposi sarcoma cells Differently, studies on several prostate and ovarian cancer cell lines evidenced that the activation of ERK signalling by the putative cannabinoid receptor GPR55 activates an autocrine loop that sustains cell proliferation It is well known that the activation of death pathways needs to be accompanied by downregulation of survival factors.
On the contrary, inhibition of pAKT leads to cell cycle arrest which precedes apoptotic response very often mediated by the involvement of intrinsic mitochondria-dependent pathway. Downregulation of AKT is involved in cannabinoid antitumoral action.
In human gastric cancer cells, it has been reported that cannabinoid receptor agonists induce cell cycle arrest which is a consequence of AKT inhibition related to the activation of MAPK pathway Interesting results were also obtained in astrocytoma In this model, cannabinoids induce apoptosis only in cells expressing low levels of CB receptors, while in cells with high CB receptor levels, cannabinoids are ineffective because of the contemporaneous high amount of the phosphorylated pro-survival AKT.
These results suggest that the coupling of CB receptors to AKT pathway when these receptors are expressed at mid and high levels eliminates the ability of cannabinoids to induce apoptosis in astrocytoma cells.
As previously indicated, autophagy can assume different and opposite roles in cell fate. In fact, this process may trigger survival pathways, collaborate with apoptosis to induce cell death or substitute itself to apoptosis to start an autonomous death pathway. Numerous recent studies have indicated the cannabinoid-mediated induction of autophagy in different experimental cancer models.
Noteworthy is the observation that the activation of autophagic pathway is often mediated by the same molecules that are involved in apoptosis. In these cells autophagy seems to be upstream of apoptosis in cannabinoid-induced cancer cell death. The stimulation of autophagy-mediated apoptosis induced by cannabinoids has also been demonstrated in animal models of cancer, including glioma 54 , Similar studies performed to investigate the effects of cannabinoids on the growth of hepatocellular carcinoma cells have demonstrated that THC markedly reduces the viability of the human hepatocellular carcinoma cell lines through induction of autophagy mediated, also in this case, by upregulation of TRB3 and subsequent inhibition of the serine-threonine kinase AKT and AMPK stimulation.
As in glioma animal model, in vivo studies on hepatocellular carcinoma subcutaneous xenografts have revealed that THC-dependent growth inhibition is reduced when autophagy is genetically or pharmacologically blocked, thus confirming that the induction of autophagy can be a key step in cannabinoid-induced cell death Further biochemical pathways have been suggested in the attempt to clarify the molecular mechanisms of cell death induced by cannabinoids.
In addition, data have also shown the activation of intrinsic apoptotic pathway. The coexistence of autophagy and apoptosis has been confirmed by different observations: A new recent study on the interplay between autophagy and apoptosis induced by cannabinoids showed a surprise in mantle cell lymphomas Interestingly, this study shows that the response to cannabinoid treatment decreases cell viability which does not involve the caspase-3 cascade.
Moreover, mantle cell lymphoma primary cells respond to cannabinoid treatment through the formation of cycloheximide-sensitive cytoplasmic vacuoles. However, the lack of enhanced autophagosome formation and lysosomal contribution also excludes the involvement of a canonical autophagic process. The authors hypothesize that the observed features resemble paraptosis-like cell death, a third type of programmed cell death, not previously described in response to cannabinoids.
Activation of more types of cell death by cannabinoids widens their potential therapeutic usefulness in cells overexpressing cannabinoid receptors. Recent studies have demonstrated the ability of cannabinoids to synergize with other molecules to trigger death pathways in cancer cells. It is well known that in cancer therapy the employment of combinations of drugs rather than a single drug represents a therapeutic strategy with distinct advantages.
On the one hand, the contemporaneous activation of different biochemical pathways can achieve synergistic effects; on the other hand, the combination can result in a reduction of the dose of each single drug thereby reducing side effects.
Thus, the ability of cannabinoids in synergizing with other drugs to improve their anticancer activity has been investigated.
In particular, we have recently demonstrated that the synthetic cannabinoid WIN55, sensitizes hepatocellular carcinoma cells to apoptosis, mediated by tumor necrosis-related apoptosis inducing ligand TRAIL. This event seems to be related to the increase in the level of p8 and CHOP, two factors implicated in ER stress response and apoptosis.
Moreover, WIN55, treatment also induces a marked downregulation of some survival factors. Therefore, both the induction of DR5 and the decrease of survival factors explain synergistic effects of the drugs in hepatocellular carcinoma cells.
Our unpublished data obtained in osteosarcoma cells seem to indicate that WIN also triggers an autophagic pathway with the increase in the level of beclin-1 and LC-3 II, but this pathway is not carried out because of the lysosomal membrane permeabilization.
Also, in pancreatic cancer cells the combination of cannabinoids with gemcitabine, a pyrimidine analog largely employed in anticancer therapy, induces synergistic effects via activation of autophagy In this case, gemcitabine induces upregulation of both CB receptors thus sensitizing cells to cannabinoid effects.
A central role in this pathway seems to be related to the increase in ROS production, induction of ER-stress which carried out to specific cell death pathway of type II autophagy. A curious observation about the therapeutic potential of cannabinoids is that in the first example cannabinoids sensitize the cells to TRAIL-induced cell death while in the latter the cannabinoid death action is potentiated by gemcitabine addition in pancreatic cancer cells.
Another example of synergistic effects of cannabinoids with other drugs has also been reported by Gustafsson et al 60 who demonstrated that the synthetic cannabinoid HU, anandamide and its other derivatives induce synergistic and cytotoxic, rather than antiproliferative, effects when employed in combination with the classic pyrimidine antagonist 5-fluorouracil 5-FU in the colorectal carcinoma cells.
The authors report that the effect does not seem to involve cannabinoid receptors and suggest the involvement of ER-stress because the employment of common antioxidants attenuates cannabinoid cytotoxicity. Autophagy is also involved in the strong antitumoral effects induced in glioma xenografts by combined administration of THC and temozolomide, the benchmark agent for the management of glioblastoma multiforme, an effect that is also observed in tumors which are resistant to temozolomide treatment.
Data reported in this review seem to confirm the ability of cannabinoids to induce cell death in different tumor models. Moreover, it can be seen from the brief literature overview presented here that these compounds are effective in inducing the main cell death modes, i. Unfortunately, to simplify the mechanisms induced by cannabinoids to carry out cells to death and discriminate the role played by each intracellular mediator appears to be very difficult for different reasons: This paper reports data derived from the Cannabinoids in MS CAMS study, which was the largest randomized controlled trial yet conducted to evaluate the therapeutic efficacy of cannabinoids.
Further work is needed to establish the functional significance of cannabinoid receptors on immune cells. Multiple sclerosis MS is the most common cause of neurological disability in young adults, with a prevalence of around one in in most of the United Kingdom [ 1 ].
While the precise aetiology of MS is unknown, there is much evidence implicating an autoimmune mechanism, with central nervous system white matter tracts providing the focus for inflammation and demyelination.
Repeated inflammatory attack eventually provokes axonal loss and glial scarring, which constitute the neuropathological basis for chronic disability and progressive disease. Treatments for MS can broadly be divided into symptomatic such as muscle relaxants and bladder treatments and so-called disease modifying, most of which are immunomodulatory, such as beta-interferons and glatiramer acetate.
Although cannabinoids have been used mainly to alleviate symptoms, there is experimental evidence to suggest that they may be immunomodulatory. We investigated the nature of potential cannabinoid immunomodulation on samples obtained from patients with MS taking part in the CAMS study, and report the results here.
Patients with stable MS were recruited into a double-blind placebo-controlled trial looking at the effect of oral cannabinoids on spasticity, the CAMS study [ 3 ]. Patients from two centres Plymouth and National Hospital, Queen Square, London provided blood samples for further laboratory analysis. The full inclusion and exclusion criteria have been published elsewhere [ 3 ], but patients were required to have had stable MS for the 6 months and no corticosteroids for 30 days prior to study entry.
Any relapses or corticosteroid medication during the study were recorded. Serum was available from patients taken before treatment and at the end of treatment, 13 weeks later for cytokine and C-reactive protein CRP measurements. Because timing of clinic attendance varied between patients, it was not possible to obtain samples at the same point after medication in all patients, although wherever possible each patient attended at the same time of day at each clinic visit.
The relatively long half-life of cannabinoids and pharmacokinetic modelling suggested that after a number of weeks patients would achieve a steady state of serum cannabinoid levels, so we would not expect short-term confounding of any longer-term effects.
Lithium heparin blood samples were taken for cell culture at visits 2 and 7. Controls were obtained from random anonymized patient samples sent to the immunology laboratory for routine immunological analyses. Peripheral blood mononuclear cells PBMC were isolated within 12 h of blood sampling see below.
CRP levels were measured as a marker of concurrent infection and a surrogate marker of IL Control wells contained monensin only. The cells were subsequently harvested, washed, fixed and permeabilized using a commercial kit Dako Intrastain; Becton-Dickinson, Oxford, UK. Measurements were taken in duplicate using samples from each of the cell culture wells. All experiments and data collection were completed prior to unblinding. The study was approved by the South-West multi-centre research ethics committee and was undertaken under licence from the UK Home Office.
All participants provided written informed consent. A total of patients were recruited to the main CAMS study from the two centres Plymouth, 79; Queen Square, 51 after the initial pilot phase of 20 patients had been recruited in Plymouth. Samples were available from patients, two of whom did not commence medication.
The disposition of patients according to treatment group and disease type is shown in Table 1. Only two patients in this cohort had a relapse treated with 1 g methylprednisolone for 3 days during the study period, one in each of the cannabis extract and placebo groups.
Grouping the two active treatment groups together gave similar results and there were no significant treatment effects on measured cytokines when the cohort was analysed according to disease type PPMS or SPMS. The asterisk represents an extreme value more than three interquartial ranges from the box hinges. Asterisks represent extreme values more than three interquartial ranges from the box hinges; open circles represent outliers more than 15 interquartial ranges from the box hinges.
Using the Mann—Whitney U -test and Wilcoxon matched pairs, there was no evidence of a significant difference in serum cytokine levels between controls and MS patients Table 3. Mann—Whitney U -test comparing serum cytokine results in vivo between MS patients and controls. Peripheral blood mononuclear cells PBMC were cultured with progressive multiple sclerosis PMS , ionomycin and monensin for 7 h, fixed and permeabilized. Results from the present experiments do not demonstrate any significant effects of cannabinoids on the cytokine profiles examined.
However, numbers are small and standard deviations large, so that relatively small but possibly clinically useful effects cannot be excluded from these results. The small numbers also make it very difficult to distinguish any differences between cytokine profiles in primary and secondary progressive disease patterns. The majority of cannabinoid actions are thought to be mediated via cannabinoid receptors, two types of which have been identified and cloned.
CB 1 receptors [ 11 ] are found predominantly in the central nervous system whereas CB 2 receptors [ 12 ] are principally located peripherally, especially on leucocytes. The majority of experimental evidence suggests that cannabinoids are anti-inflammatory and may be associated with a Th1 to Th2 shift. This may be mediated via a reduction in IL [ 14 ]. The correlation between serum level and symptomatic or immunological effect is the focus of ongoing work.
Cannabinoid-associated cell death mechanisms in tumor models (Review)
Request PDF on ResearchGate | Cannabinoids, Immune System and Cytokine Network | How cannabinoids influence immune function has been examined. Jul 31, Request PDF on ResearchGate | Interaction between cytokines, of the cannabinoid system is associated with therapeutic effects that may be. CANNABINOID EFFECTS ON. IMMUNE FUNCTION. The immune system is a complex network of cells, tissues and soluble mediators, including cytokines and .