Wow! Does Cannabis has some awesome uses or what? Talk about save your bone when your sick! This cannabis extract is going to put the hurt on alot of pharma companies that just want your REPEAT business.
Lets Learn about TB and the phases… Below is a wiki clip to better educate your self.
Below that is the medical report of the Effects of CBD on TB. Kinda confusing and hard to understand. But do your best to understand doctor lingo :)
If you suffer currently, or are in the latent phase of TB, GET SOME CBD to start healing now!
Please share this post with anyone that has to deal with pharma.
—————————————————————————-The Spirit Smoker
Tuberculosis, MTB, or TB (short for tubercle bacillus), in the past also called phthisis, phthisis pulmonalis, or consumption, is a widespread, and in many cases fatal, infectious disease caused by various strains of mycobacteria, usually Mycobacterium tuberculosis. Tuberculosis typically attacks the lungs, but can also affect other parts of the body. It is spread through the air when people who have an active TB infection cough, sneeze, or otherwise transmit respiratory fluids through the air. Most infections do not have symptoms, known as latent tuberculosis. About one in ten latent infections eventually progresses to active disease which, if left untreated, kills more than 50% of those so infected.
The classic symptoms of active TB infection are a chronic cough with blood-tinged sputum, fever, night sweats, and weight loss (the latter giving rise to the formerly common term for the disease, “consumption”). Infection of other organs causes a wide range of symptoms. Diagnosis of active TB relies on radiology (commonly chest X-rays), as well as microscopic examination and microbiological culture of body fluids. Diagnosis of latent TB relies on the tuberculin skin test (TST) and/or blood tests. Treatment is difficult and requires administration of multiple antibiotics over a long period of time. Social contacts are also screened and treated if necessary. Antibiotic resistance is a growing problem in multiple drug-resistant tuberculosis (MDR-TB) infections. Prevention relies on screening programs and vaccination with the bacillus Calmette-Guérin vaccine.
BACKGROUND AND PURPOSE
Cannabis extracts and several cannabinoids have been shown to exert broad anti-inflammatory activities in experimental models of inflammatory CNS degenerative diseases. Clinical use of many cannabinoids is limited by their psychotropic effects. However, phytocannabinoids like cannabidiol (CBD), devoid of psychoactive activity, are, potentially, safe and effective alternatives for alleviating neuroinflammation and neurodegeneration.
We used experimental autoimmune encephalomyelitis (EAE) induced by myelin oligodendrocyte glycoprotein (MOG) in C57BL/6 mice, as a model of multiple sclerosis. Using immunocytochemistry and cell proliferation assays we evaluated the effects of CBD on microglial activation in MOG-immunized animals and on MOG-specific T-cell proliferation.
Treatment with CBD during disease onset ameliorated the severity of the clinical signs of EAE. This effect of CBD was accompanied by diminished axonal damage and inflammation as well as microglial activation and T-cell recruitment in the spinal cord of MOG-injected mice. Moreover, CBD inhibited MOG-induced T-cell proliferationin vitro at both low and high concentrations of the myelin antigen. This effect was not mediated via the known cannabinoid CB1 and CB2 receptors.
CONCLUSIONS AND IMPLICATIONS
CBD, a non-psychoactive cannabinoid, ameliorates clinical signs of EAE in mice, immunized against MOG. Suppression of microglial activity and T-cell proliferation by CBD appeared to contribute to these beneficial effects.
The anti-inflammatory properties of cannabinoids, constituents of the Cannabis sativa plant, have been appreciated since ancient times and have been supported experimentally, in models of inflammation in vitro and in vivo (Tanasescu and Constantinescu, 2010). Indeed, immune cells express the elements of the cannabinoid system including endocannabinoid ligands, endocannabinoid enzymatic machinery and cannabinoid CB receptors, mostly of the CB2 type and many fewer of the CB1 type (Mackie, 2005; nomenclature follows Alexander et al. (2009). Moreover, expression of CB1 and CB2 receptors and the activity of the endocannabinoid system are regulated in response to inflammation, a finding that further confirms the involvement of the cannabinoid system in immune processes (Croxford and Yamamura, 2005; Maresz et al., 2005; Pietr et al., 2009).
The clear and promising therapeutic potential of cannabinoids (Klein and Newton, 2007) is limited due to central, CB1-mediated, psychotropic effects of many of these materials. A good example is Δ9-tetrahydrocannabinol (THC) as the most studied psychoactive cannabinoid, activating both CB1 and CB2 receptors (Bayewitch et al., 1995). The preferential CB2 receptor expression on immune cells offers an attractive opportunity to regulate the function of the immune system with CB2 receptor ligands. Studies on CB1/CB2 receptor knock-down mice revealed the existence of other possible, receptor and non-receptor, cannabinoid targets (Járai et al., 1999), confirmed to be immunomodulatory (Kaplan et al., 2003; 2008;). Thus, cannabinoid compounds not acting on CB1 or CB2 receptors seem to offer new tools to manipulate inflammation. Cannabidiol (CBD) is one of these compounds and is the major Cannabis-derived non-CB1/CB2 receptor ligand (Showalter et al., 1996). This compound is not only devoid of psychotropic effects but also is able to inhibit many central effects of CB1receptor ligands, for example, the anxiogenic and psychotogenic activities of THC (Zuardi, 2008). Interestingly, despite different pharmacological and behavioural effects, CBD shares with ‘classic’ psychocannabinoids many beneficial effects, including its capacity to act as an immunomodulator.
Indeed, CBD exerts a wide range of anti-inflammatory properties and regulates cell cycle and function of various immune cells. These effects include suppression of humoral responses, such as release of cytokines, chemokines, growth factors, as well as suppression of immune cell proliferation, activation, maturation, migration and antigen presentation (Mechoulam et al., 2007). In an earlier publication, we showed that CBD inhibited production of the cytokine IL-6 and the chemokine CCL-2 by activated microglial cells and in parallel activated intracellular anti-inflammatory pathways (Kozela et al., 2010).
Among the many types of neurodegenerative diseases in which inflammation is involved, multiple sclerosis (MS) is one of those clearly induced and driven by dysfunctional immune system activity. In MS, myelin autoreactive peripheral T cells migrate into the CNS and initiate cytotoxic, degenerative processes that include demyelination, oligodendrocyte cell death and axonal degeneration. These effects lead to neurological deficits such as visual and sensory disturbances, motor weakness, tremor, ataxia and progressive disability as the main clinical symptoms (Compston and Coles, 2008). Infiltrating T cells are constantly reactivated within the CNS parenchyma by microglia. Microglial cells via chemokine and cytokine release and constant antigen presentation potentiate T-cell recruitment to the CNS and facilitate their polarization into cytotoxic phenotypes (Th1 or Th17). Moreover, via released chemokines such as CCL-2, microglial cells recruit other immune cells of myeloid origin, specialized in epitope spreading and phagocytosis of myelin including monocytes, macrophages, B cells and dendritic cells (Jacket al., 2005; Koning et al., 2009). Depletion of microglia or impairment of their function can attenuate disease progression in experimental animal models of MS supporting their role in initiation and development of this disease (Huitinga et al., 1990; Heppner et al., 2005). Thus, suppression of microglia will potentially reduce inflammatory lesions and limit demyelination within the CNS.
One of the best described and commonly used animal models of MS is experimental autoimmune encephalomyelitis (EAE) which is induced when the animals are immunized with myelin components, for example, myelin basic protein, proteolipid protein or myelin oligodendrocyte glycoprotein (MOG) or by passive transfer of autoreactive myelin specific T cells to produce demyelination and MS-like neurological and clinical signs (Shevach, 1999). Using the MOG-induced EAE mouse model, we investigated if systemically given CBD at the time of symptomatic disease onset could affect the progression of the disease. We observed that CBD ameliorated the severity of the EAE in MOG-injected mice. Moreover, CBD attenuated microglia activation in MOG-immunized animals and inhibited MOG-induced proliferation of encephalitogenic T cells.
Animals and experimental design
All animal care and experimental procedures complied with and were approved under the guidance and regulations of the Weizmann Institute of Science and Tel Aviv University. EAE was induced in 30 8-week-old female C57BL/6 mice (Harlan Laboratories, Rehovot, Israel) by two subcutaneous injections of MOG35-55 fragment (encompassing amino acids 35–55 of MOG) on days 1 and 8, injected in the left and right flanks respectively. Each injection contained 300 µg MOG in 200 µL of complete Freund’s adjuvant (CFA) containingMycobacterium tuberculosis at 200 µg (Sigma, Israel). Control mice received only CFA without MOG (Ctrl, n= 10).
CBD (5 mg·kg−1; CBD + EAE group) or its vehicle (cremophor, ethanol and PBS, v : v : v 1:1:18; EAE group) was injected (i.p.) into 15 MOG-immunized mice immediately at the onset of disease signs for 3 consecutive days (on days 19, 20 and 21). Clinical disease scores were recorded daily until 30 days after first immunization. At this time point, spinal cords were collected for further pathological and immunological studies. The signs of EAE were scored as follows: 0, no clinical signs; 1, loss of tail tonicity; 2, partial hind limb paralysis; 3, complete hind limb paralysis; 4, partial frontal limb paralysis; 5, total paralysis; 6, death (Lev et al., 2004). In parallel, we used control mice which received only CFA (Ctrl) and were also injected with CBD on days 19, 20 and 21 (Ctrl + CBD group, each group n = 10). The days of EAE onset and respective CBD injections were chosen based on our previous experience with this model (Lev et al., 2004).
CBD (National Institute on Drug Abuse, Rockville, MD, USA) solution was prepared freshly before each of the treatments. The dose of CBD was chosen based on previous studies with systemic administration of the drug in which CBD at 5 mg·kg−1 exhibited maximal efficiency at relieving rheumatoid signs in collagen-induced arthritis (Malfait et al., 2000). MOG synthesis was carried out by the Weizmann Institute Synthesis Unit, using a solid-phase technique on a peptide synthesizer (Applied Biosystems Inc., Foster City, CA, USA).
Histology and immunocytochemistry
Histological analysis of spinal cord sections was used to define severity of inflammation and demyelination and the effect of CBD treatment on these parameters. Spinal cords were dissected 30 days after first immunization of mice, fixed in 10% buffered formalin and embedded in paraffin. The presence of axonal pathology was supported by immunohistochemistry with anti-non-phosphorylated neurofilament H (SMI-32, Sternberg Antibodies, Emeryville, CA, USA) on 8 µm thick paraffin sections. Subsequently, 4 µm thick sections were stained with haematoxylin and eosin (H&E) and assessed for inflamed areas. The assessment of immune cell infiltration was performed using immunocytochemistry on 4 µm spinal cord sections and included staining for T cells (CD3+, Serotec, Kiddlington, Oxford, UK), microglia/macrophage number and their activation state (Iba-1, Wako, Richmond, VA, USA; and Mac-2/Galectin 3, Cedarlane, Burlington, Ontario, Canada).
It should be noted that the proteins, ionizing calcium-binding adaptor molecule 1 (Iba-1) and Mac-2/Galectin-3, do not discriminate between microglia, resident macrophages within the CNS, and invasive macrophage-like cells from peripheral sites that crossed the blood-brain barrier. Thus, in further description of the stainings we refer to microglia/macrophage populations. The Iba-1 protein is specifically expressed in microglia/macrophages and becomes up-regulated during the activation of these cells. Mac-2/Galectin-3 is another microglial/macrophage marker whose expression reflects specifically the activation state of these cells. For example, Mac-2/Galectin-3 expression is known to be increased in these cells following phagocytosis of damaged myelin and cell particles (Reichert and Rotshenker, 1999).
The immunostainings were visualized with respective secondary antibodies conjugated to Cy2 or Cy3 fluorochromes. Spinal cords from control, healthy mice that received 3 daily injections of either CBD or its vehicle were dissected and stained in parallel. Assessments of intensity and differences in immunofluorescence staining for Iba-1 and Mac-2/Galectin-3 were performed using the Image Pro Plus analysis software (Media cybernetics, Bethesda, MD, USA) and collected as arbitrary units representing optical density per area. The statistics was carried out on per cent values calculated from optical density values and is given in the appropriate figure legend. In the case of T cells, CD3+ cells were counted (three areas per mouse, three to four mice per treatment) and the cell numbers were subjected to statistical analysis.
Encephalitogenic T-cell line
The MOG35-55-specific T-cell line was established from lymph node cells of C57BL/6 female mice that had been primed 10 days earlier with MOG35-55 emulsified in CFA as previously described (Kaushansky et al., 2006). The T-cell line was maintained in vitro in RPMI-1640 (Biological Industries, Kibbutz Beit HaEmek, Israel) containing 5% fetal calf serum and supplemented with 10 U·mL−1 IL-2 (Peprotech Inc, Rocky Hill, NJ, USA), L-glutamine, 100 µg·mL−1 of streptomycin, 100 U·mL−1 of penicillin, 50 µM β-mercaptoethanol, 1 mM non-essential amino acids and 1 mM sodium pyruvate with alternate stimulation with MOG every 14 days as previously described (Ben-Nun and Lando, 1983; Kaushansky et al., 2006).
Encephalitogenic T-cell proliferation
The MOG35-55-reactive line of T cells raised from primed mice was cultured in 96-well plates (1.5 × 104cells·per well) together with irradiated (25Gy) splenic antigen presenting cells (APCs; 5 × 105 cells per well). APCs were isolated from spleens of naïve C57BL/6 mice. Lysis of erythrocytes was performed using ACK solution (150 mM NH4Cl, 10 mM KHCO3 and 0.1 mM EDTA). The cells were cultured in 0.2 mL RPMI-1640 containing 2.5% fetal calf serum and supplemented with L-glutamine, 50 µM β-mercaptoethanol, and in the presence of MOG35-55 peptide (1 or 2.5 µg·mL−1). CBD at final concentrations of 1, 5 or 10 µM or its vehicle (0.1% ethanol in RPMI-1640) was added to the cells together with the MOG. The cells were then incubated for 72 h at 37°C in 5% CO2 humidified air (Ben-Nun and Lando, 1983; Kaushansky et al., 2006). Cell proliferation was measured by pulsing the cells with [3H]Thymidine (0.5 µCi·well−1) for the last 16 h of the incubation period and the cells were then harvested and counted using a Matrix 96 Direct β counter (Packard Instr., Meriden, CT, USA). To evaluate if CB1 or CB2 receptors were involved in the effects of CBD on T-cell proliferation, SR141716 (CB1 receptor antagonist; RTI International, NC, USA) and SR144528 (CB2 receptor antagonist; RTI International, NC, USA) at 1 µM concentration were applied 30 min before the application of CBD (5 or 10 µM) and MOG at 1 µg·mL−1. The proliferative responses of the cell microcultures were converted to per cent values with MOG effect expressed as 100%. Per cent values were calculated from Stimulation Index (SI) values which are the fold changes in mean counts per minute (cpm) of MOG cultures over mean cpm of cultures without MOG (spontaneous proliferation). Statistical analysis was performed on per cent values. Each experiment was carried out in triplicates and repeated three or four times.
EAE clinical scores were reported as average ± SD. Repeated measure analysis of variance (anova) was used for statistical analysis of these data. The fluorescence intensity and T-cell proliferation data were expressed as the mean ± SEM and analysed for statistical significance using one-way anova, followed by Bonferroni post hoctests. P < 0.05 was considered significant. Graph Pad Prism program (La Jolla, CA, USA) was used for statistical analysis of the data.
CBD ameliorates clinical signs of MOG-induced EAE and disease progression
C57BL/6 mice immunized with MOG35-55 developed severe EAE disease with complete hind limb paralysis [EAE mice, average clinical score 2.52 ± 0.54; Figure 1; with 13 out of 15 mice exhibiting clinical signs of EAE (EAE ratio 13/15)]. As shown in Figure 1, three injections (i.p.) of CBD (5 mg·kg−1, one a day) during the onset of clinical disease resulted in amelioration of the disease signs during the days of injections as well as markedly delaying disease progression. On day 21, MOG-induced EAE mice exhibited limp tail and partial paresis of the hind limbs (clinical score 1.03 ± 0.29; EAE ratio 9/15). However, MOG-induced EAE mice that received CBD during days 19–21 exhibited minimal clinical signs of EAE (average clinical score 0.13 ± 0.09, P < 0.05; Figure 1; EAE ratio 1/15). Interestingly, 2 days after the CBD injections (day 23), the clinical score of CBD-treated mice increased slowly, reaching mild disease severity at the end of the study (average clinical score 1 and EAE ratio 6/15 vs. average clinical score of 2.54 ± 0.54 in untreated EAE mice, P < 0.05; EAE ratio 13/15).
Consequently, CBD, a compound active on non-CB1 or CB2 receptors (Showalter et al., 1996), has been shown to possess a wide anti-inflammatory profile. CBD was shown to decrease TNFα, IL-2 and IFNγ release from activated splenocytes and macrophages (Malfait et al., 2000; Jan et al., 2007; Kaplan et al., 2008). It also suppresses concavalin A and collagen induced T-cell proliferation (Malfait et al., 2000; Jan et al., 2007), microglial migration (Walter et al., 2003) and cytokine release (Kozela et al., 2010). It suppresses antigen-specific antibody production in splenocytes (Jan et al., 2007), as well as attenuates endothelial inflammation and barrier disruption (Rajesh et al., 2007). These activities of CBD may contribute to its beneficial effects in EAE, as observed in our hands, because many of these immune processes were reported to be involved in EAE pathology at different stages of the disease model.
Anti-oxidant and neuroprotective properties of CBD
Several cannabinoids including THC and CBD exert anti-oxidative and neuroprotective properties (Mechoulam et al., 2007). Most of the current MS therapies are directed against various immune cells to achieve immunosuppressive effects. However, increasing evidence shows that immunosuppression alone was not sufficient for therapeutic effect especially in late, secondary progressive MS (Bennett and Stüve, 2009; Jones and Coles, 2010). In these cases, the neurodegenerative processes become resistant to immunomodulation. Indeed, neurodegeneration that consists of neuronal and axonal loss can result from oxidative stress and excitotoxicity and is driven by activated microglial cells and macrophagic/monocytic infiltrates (Hanisch and Kettenmann, 2007). It appears that the cannabinoid system could provide a rescue mechanism in such conditions. Accordingly, the ameliorating activity of THC-like cannabinoids combines CB2 receptor-mediated inhibition of autoreactive T cells and CB1 receptor-mediated neuroprotective activity on neurons (Maresz et al., 2007). Similarly, Croxford et al. (2008) pointed out that cannabinoid-mediated neuroprotection rather than immunosuppression, was relevant for the recovery process at the later, remissive stages of MS.