METHODOLOGY/PRINCIPAL FINDINGS: The cytotoxic effect of thymoquinone was assessed using an MTT assay, while the inhibitory effect of thymoquinone on murine WEHI-3 cell growth was due to the induction of apoptosis, as evidenced by chromatin condensation dye, Hoechst 33342 and acridine orange/propidium iodide fluorescent staining. In addition, Annexin V staining for early apoptosis was performed using flowcytometric analysis. Apoptosis was found to be associated with the cell cycle arrest at the S phase. Expression of Bax, Bcl2 and HSP 70 proteins were observed by western blotting. The effects of thymoquinone on BALB/c mice injected with WEHI-3 cells were indicated by the decrease in the body, spleen and liver weights of the animal, as compared to the control.
CONCLUSION: Thymoquinone promoted natural killer cell activities. This compound showed high toxicity against WEHI-3 cell line which was confirmed by an increase of the early apoptosis, followed by up-regulation of the anti-apoptotic protein, Bcl2, and down-regulation of the apoptotic protein, Bax. On the other hand, high reduction of the spleen and liver weight, and significant histopathology study of spleen and liver confirmed that thymoquinone inhibited WEHI-3 growth in the BALB/c mice. Results from this study highlight the potential of thymoquinone to be developed as an anti-leukemic agent.
HYPOTHESIS/ PURPOSE: To compare the anti-inflammatory activities and the anti-nociceptive properties of RG and BG.
METHODS: Nitric Oxide (NO) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) assay, quantitative Reverse Transcriptase-Polymerase Chain Reaction (qRT-PCR), western blot, xylene-induced ear edema, carrageenan-induced paw edema RESULTS: The ginsenoside contents were confirmed using high-performance liquid chromatography (HPLC) and has been altered through increased processing. The highest concentration of these extracts inhibited NO production to near-basal levels in lipopolysaccharide (LPS)-stimulated RAW 264.7 without exhibiting cytotoxicity. Pro-inflammatory cytokine expression at the mRNA level was investigated using qRT-PCR. Comparatively, BG exhibited better inhibition of pro-inflammatory mediators, iNOS and COX-2 and pro-inflammatory cytokines, IL-1β, IL-6 and TNF-α. Protein expression was determined using western blot analysis and BG exhibited stronger inhibition. Xylene-induced ear edema model in mice and carrageenan-induced paw edema in rats were carried out and tested with the effects of ginseng as well as dexamethasone and indomethacin - commonly used drugs. BG is a more potent anti-inflammatory agent, possesses anti-nociceptive properties, and has a strong potency comparable to the NSAIDs.
CONCLUSION: BG has more potent anti-inflammatory and anti-nociceptive effects due to the change in ginsenoside component with increased processing.
PURPOSE: In the present study, phyllanthin isolated from Phyllanthus amarus was investigated for its immunosuppressive effects on various cellular and humoral immune responses in Balb/C mice.
METHODS: Male mice were treated daily at 20, 40 and 100mg/kg of phyllanthin for 14 days by oral gavage. The effects of phyllanthin on cellular immune responses in treated /non treated mice were determined by measuring CD 11b/CD 18 integrin expression, phagocytosis, nitric oxide (NO) production, myeloperoxidase activity (MPO), T and B cells proliferation, lymphocyte phenotyping, serum cytokines production by activated T-cells and delayed type hypersensitivity (DTH). Its effects on humoral immune responses were evaluated by determining the serum levels of lysozyme and ceruloplasmin, and immunoglobulins (IgG and IgM).
RESULTS: Phyllanthin dose-dependently inhibited CD11b/CD18 adhesion, the engulfment of E. coli by peritoneal macrophages molecules, NO and MPO release in treated mice. Phyllanthin caused significant and dose-dependent inhibition of T and B lymphocytes proliferation and down-regulation of the Th1 (IL-2 and IFN-γ) and Th2 (IL-4) cytokines. Phyllanthin at 100mg/kg caused a significant reduction in the percentage expression of CD4(+) and CD8(+) in splenocytes and the inhibition was comparable to that of cyclosporin A at 50mg/kg. At 100mg/kg, phyllanthin also dose-dependently exhibited strong inhibition on the sheep red blood cell (sRBC)-induced swelling rate of mice paw in DTH. Significant inhibition of serum levels of ceruloplasmin and lysozyme were observed in mice fed with higher doses (40 and 100mg/kg) of phyllanthin. Anti-sRBC immunoglobulins (IgM and IgG) antibody titer was down-regulated in immunized and phyllanthin-treated mice in a dose-dependent manner with maximum inhibition being observed at 100mg/kg.
CONCLUSION: The strong inhibitory effects of phyllanthin on the cellular and humoral immune responses suggest that phyllanthin may be a good candidate for development into an effective immunosuppressive agent.
METHODS: A total of six conserved peptides representing B- and T-cell epitopes of Influenza A were identified and they were formulated in either incomplete Freund's adjuvant containing CpG ODN 1826 or being encapsulated in PLGA nanoparticles for the evaluation of immunogenicity in BALB/c mice.
RESULTS: The self-adjuvanting PLGA nanoparticles encapsulating the six conserved peptides were capable of eliciting the highest levels of IgG and IFN- γ producing cells. In addition, the immunogenicity of the six peptides encapsulated in PLGA nanoparticles showed greater humoral and cellular mediated immune responses elicited by the mixture of six naked peptides formulated in incomplete Freund's adjuvant containing CpG ODN 1826 in the immunized mice. Peptide 3 from the mixture of six peptides was found to exert necrotic effect on CD3+ T-cells and this finding indicated that peptide 3 should be removed from the nanovaccine formulation.
CONCLUSION: The study demonstrated the self-adjuvanting properties of the PLGA nanoparticles as a delivery system without the need for incorporation of toxic and costly conventional adjuvants in multi-epitope peptide-based vaccines.