Beta-caryophyllene as an antioxidant, anti-inflammatory and re-epithelialization activities in a rat skin wound excision model.

The skin is a critical organ for the maintenance of the integrity and protection of the organism. When a wound occurs, a sequence of healing mechanisms is triggered to reconstruct the wounded area. ß-caryophyllene is a sesquiterpene in Copaifera langsdorffii oleoresin with antioxidant and anti-inflammatory potential. On the basis of previous studies with C. langsdorffii, ß-caryophyllene was selected to evaluate its wound healing potential and pharmacological mechanisms. The excision wound model was used with male Wistar rats and macroscopic, histological, immunohistochemical and biochemical analyses were performed with skin samples, comparing the ß-caryophyllene-treated group with reference drugs. The results showed macroscopic retraction of the wounds treated with ß-caryophyllene. Biochemical assays revealed the antioxidant and anti-inflammatory mechanisms of the ß-caryophyllene-treated group with increasing levels of IL-10 and GPx and decreasing levels of pro-inflammatory molecules, including TNF-α, IFN-γ, IL-1ß and IL-6. After ß-caryophyllene treatment, immunohistochemical assays showed enhanced re-epithelialization, through the increase in laminin-γ2 and desmoglein-3 immunolabeling. ß-caryophyllene also act in the remodeling mechanism, increasing the collagen content in the Masson's trichrome staining. These findings indicated the wound-healing potential of ß-caryophyllene topical formulation in rat skin wounds, mediated by antioxidant, anti-inflammatory and re-epithelialization mechanisms.

Therapeutic effects of ß-caryophyllene on proliferative disorders and inflammation of the gerbil prostate.

BACKGROUND: The prostate is susceptible to changes in androgen levels, which can play an important role in the development of Benign Prostatic Hyperplasia (BPH). Natural compounds have beneficial properties for organisms and can be an important therapeutic strategy in the treatment of diseases. ß-Caryophyllene (BCP) is a phytocannabinoid present in several medicinal and food plants species and has shown beneficial effects in different organs. However, little is known about its effects on the prostate. The present study seeks to evaluate the effects of exposure to BCP on the morphophysiology of the ventral prostate of adult gerbils supplemented with testosterone. METHODS: Animals were distributed into four groups (n = 8/group): Intact control (C); ß-Caryophyllene (BCP): ß-Caryophyllene (50 mg/kg/day); Testosterone (T): animals received subcutaneous injections of Testosterone Cypionate (3 mg/Kg), on alternate days, for one month and were euthanized 30 days supplementation ended; Testosterone and ß-Caryophyllene (TBCP): animals were exposed to testosterone cypionate (3 mg/Kg) to induce hyperplastic alterations followed by daily BCP (50 mg/kg). Morphological, biometric, immunohistochemical, and serological analyses were performed. RESULTS: Proliferative disorders and inflammatory foci were present in the ventral prostate of all experimental groups. An increase in the multiplicity of benign intraepithelial neoplasm and subepithelial inflammatory foci was observed in T group. The incidence of intraluminal inflammatory foci and microinvasive carcinoma was verified only in the T group. Cellular rearrangement and tissue remodeling occurred in the prostate of groups exposed to phytocannabinoids. A reduction was observed in the frequency of PHH3 and Cox2 markers in the prostatic epithelium of TBCP in comparison with T. A decrease in F4/80 and CD163 positive macrophages were also observed in the prostatic stroma of the TBCP group in comparison with T. The results suggest that BCP had favorable effects on BPH, reducing the proliferation and frequency of some inflammatory cells. CONCLUSION: BCP impacts the tissue remodeling process in the premalignant prostate environment and that the use of this phytocannabinoid can have a promising effect in the handling of BPH.

Cedrol attenuates collagen-induced arthritis in mice and modulates the inflammatory response in LPS-mediated fibroblast-like synoviocytes.

Ginger has been used as a flavoring agent and traditional medicine for a long time in Asian countries. Pharmacological studies showed that it has antiemetic, anti-inflammatory and analgesic effects, which is attributed to its chemical constituents. The aim of the present study is to investigate the anti-rheumatoid arthritis properties of cedrol (CE) found in ginger. In an in vivo anti-RA study, CE remarkably alleviated the paw swelling and arthritis score in CE-treated CIA mice compared with the model group. The neutrophil count and the productions of TNF-α and IL-1ß were inhibited, and the expressions of Rankl, Cox-1 and Cox-2 were down-regulated at the mRNA level. Radiologic evaluation, histopathological analysis and immunohistochemistry indicated that CE ameliorated inflammatory cell infiltration, synovial hyperplasia, and bone and cartilage damage, and exhibited an immunotherapeutic effect. The MTT assay demonstrated that CE (10-10-10-5 M) had no cytotoxicity on fibroblast-like synoviocytes (FLSs), and exhibited an inhibitory effect on the proliferation of LPS-induced FLSs at concentrations of 10-6 M and 10-5 M. Mechanism research showed that the suppressed expressions of pivotal inflammatory mediators including COX-1 and COX-2 subsequently reduced the production of PGE2, thereby causing the secretions of pro-inflammatory cytokines, ultimately attenuating the progression of inflammation. Meanwhile, the reduction in the mRNA levels of Mmp-13 and Mcp-1 responsible for osteoclastogenesis resistance was detected. This illustrated that CE showed anti-rheumatoid arthritis properties via blocking the phosphorylation of ERK/MAPK and p65/NF-κB signaling pathways in LPS-activated FLSs. The current research suggested that CE is an important functional component in ginger, which may be a promising candidate drug for RA therapy.

Role of ß-Caryophyllene in the Antinociceptive and Anti-Inflammatory Effects of Tagetes lucida Cav. Essential Oil.

Tagetes lucida Cav. (Asteraceae) is an ancient medicinal plant commonly used to alleviate pain. Nevertheless, scientific studies validating this property are lacking in the literature. Animal models of pain were used to evaluate the antinociceptive and anti-inflammatory activities of T. lucida essential oil (TLEO) and a bioactive metabolite. The chemical constitution and possible toxicity of the extract and the mechanism of action of ß-caryophyllene were also explored. Temporal course curves and dose-response graphics were generated using TLEO (0.1-10 mg/kg or 3.16-31.62 mg/kg) and ß-caryophyllene (3.16-10 mg/kg). Metamizole (80 mg/kg) and indomethacin (20 mg/kg) were used as reference drugs in the formalin assay and writhing test in rats and mice, respectively. The ß-caryophyllene mechanism of action was explored in the presence of naloxone (1 mg/kg), flumazenil (10 mg/kg), WAY100635 (0.16 mg/kg), or nitro-l-arginine methyl ester (L-NAME) (20 mg/kg) in the formalin test in rats. GC/MS analysis demonstrated the presence of geranyl acetate (49.89%), geraniol (7.92%), and ß-caryophyllene (6.27%). Significant and dose-dependent antinociceptive response was produced by TLEO and ß-caryophyllene without the presence of gastric damage. In conclusion, ß-caryophyllene was confirmed as a bioactive compound in the T. lucida analgesic properties by involving the participation of receptors like opioids, benzodiazepines, and Serotonin 1A receptor (5-HT1A), as well as nitric oxide.

Himachalol induces apoptosis in B16-F10 murine melanoma cells and protects against skin carcinogenesis.

ETHNOPHARMACOLOGICAL RELEVANCE: Cedrus libani A. Rich (C. libani) is majestic evergreen Mediterranean conifer growing in the mountains of Lebanon. The ethnobotanical and traditional uses of cedar wood oil traces back to ancient times for the treatment of various ailments including cancer. Previous work in our laboratories revealed that himachalol (7-HC), a major sesquiterpene isolated from C. libani, possesses potent cytotoxic activity against various human cancer cell lines as well as promising anti-inflammatory effect in isolated rat monocytes. AIM OF THE STUDY: The present study aims to elucidate the mechanism of action behind the cytotoxic activity of 7-HC against murine melanoma cells (B16F-10) and evaluates its chemopreventive effect against chemically-induced skin carcinogenesis in mice. MATERIALS AND METHODS: 7-HC was extracted and purified from Cedrus libani wood. Cell viability was evaluated using WST-1 kit. Cell cycle analysis and apoptosis were assessed by Flow cytometry using propidium iodide (PI) and fluorescein Isothiocyanate (FITC)-conjugated Annexin V/PI staining respectively. Apoptosis related protein were quantified using western blot. The chemopreventive activity of 7-HC was evaluated for 20 weeks using a DMBA/TPA induced skin carcinogenesis model in Balb/c mice. RESULTS: 7-HC displayed a potent anti-proliferative activity against the melanoma cells with an IC50 of 8.8 µg/ml and 7.3 µg/ml at 24 and 48 h, respectively. Co-treatment with Cisplatin did not show any synergistic or additive effect on cell viability. Flow cytometry analysis using PI revealed that 7-HC treatment (5 and 10 µg/ml) induces the accumulation of cells in the sub-G1 phase and causes a decline in cell populations in the S and G2/M phases. Annexin/PI staining also reveals that 7-HC treatment significantly increases the percentage of cells undergoing early and late apoptosis. Western blot analysis shows that 7-HC treatment decreases the level of the anti-apoptotic protein Bcl-2 and increases the level of the pro-apoptotic protein Bax. A reduction in the level of phosphorylated Erk and Akt was also observed. 7-HC via topical (2.5%), intraperitoneal (10, 25 and 50 mg/kg) or gavage (50 mg/kg) treatment revealed a significant decrease in papilloma volume with no adverse effect on liver and kidney function. CONCLUSIONS: The present study demonstrates that 7-HC treatment protects against chemically-induced skin carcinogenesis, promotes cell cycle arrest and induces apoptosis partially through an inhibition of both the MAPK/Erk and PI3K/Akt pathways.

Nootkatone, an AMPK activator derived from grapefruit, inhibits KRAS downstream pathway and sensitizes non-small-cell lung cancer A549 cells to adriamycin.

BACKGROUND: Non-small-cell lung cancer (NSCLC) accounts for approximately 85% of all lung cancer cases and it is intrinsically resistant to anticancer drugs. Nootkatone (NKT), which is the main fragrant component of grapefruit, has been identified as a bioactive compound with a wide range of beneficial applications. NKT can activate AMP-activated protein kinase (AMPK) in liver and muscle cells, however, little is known about the role of NKT in cancer, particularly its role in NSCLC with high rates of liver kinase B1 (LKB1) and KRAS mutations. PURPOSE: The anti-cancer activities of NKT in NSCLC A549 cells and ADR-resistant A549/ADR cells were investigated and compared to those of metformin, an AMPK activator that is used clinically as an AMPK activator. METHODS: Cell viability, proliferation and NKT sensitization were determined by the MTT assay. Mechanisms of NKT against anti-cancer activities including AMPK activation, cell cycle arrest, and synergistic cytotoxic effect were evaluated by Western blot analysis, and flow cytometry. In in vivo experiments, athymic BALB/c male nude mice were used for experiments. After the successful generation of tumor models through subcutaneous injection of A549/ADR cells, NKT and/or ADR were administered and mice were kept for weekly measurements for up to 7 weeks. The animals were then sacrificed, and the tumors were removed from all animals and weighed. RESULTS: NKT activated AMPK via LKB1-independent and CAMKK2-dependent pathways, leading to inhibition of cell growth and induction of G1 cell arrest. The effect of NKT is comparable but superior to that of metformin, an AMPK activator in clinical use. Importantly, NKT inhibited the activation of oncogenic AKT and ERK proteins, while metformin inhibited AKT but failed to impact ERK, the major oncogenic protein of NSCLC cells with KRAS mutation. The synergistic activity of NKT and ADR was more effective than that of metformin and ADR. In vivo data confirmed synergistic effects of NKT and ADR without systemic side effects. CONCLUSION: We demonstrate for the first time that NKT can sensitize ADR-resistant A549/ADR cells to ADR in vitro and in vivo. Metformin, on the other hand, failed to show any synergistic effect with ADR in A549/ADR cells.