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1.
Figure 2

Figure 2. From: Reduction of human monocytic cell neurotoxicity and cytokine secretion by ligands of the cannabinoid-type CB2 receptor.

An example of immunoblot showing CB2 receptor-specific bands in extracts of human monocytic THP-1 and promyelocytic HL-60 cells. Equivalent amounts of protein (45 μg/lane) were separated through 15% polyacrylamide gels and visualized by a chemiluminescence assay. Location of molecular size markers (kDa) is shown in the left lane.

Andis Klegeris, et al. Br J Pharmacol. 2003 June;139(4):775-786.
2.
Figure 3

Figure 3. From: Reduction of human monocytic cell neurotoxicity and cytokine secretion by ligands of the cannabinoid-type CB2 receptor.

Immunostaining of cultures from surgically resected human temporal lobe tissue. (a) microglial cell stained with an antibody to CD68; (b) staining of microglia with an antibody to the CB2 receptor; (c) staining of one of a small number of astrocytes that appeared in the cultures with an antisera to GFAP. Calibration bar in (c) is for all three panels.

Andis Klegeris, et al. Br J Pharmacol. 2003 June;139(4):775-786.
3.
Figure 1

Figure 1. From: Reduction of human monocytic cell neurotoxicity and cytokine secretion by ligands of the cannabinoid-type CB2 receptor.

Polaroid photographs of a typical ethidium bromide-stained gel demonstrating PCR products for CB1 and CB2 receptors after 35 amplification cycles. Amplification products are shown for human THP-1 monocytic cells, human SH-SY5Y neuroblastoma cells, human postmortem microglia and human promyelocytic HL-60 cells. Location of molecular size markers in base pairs is shown in the left lane. Notice that CB1 receptor mRNA bands are observed in human monocytic THP-1 and SH-SY5Y neuronal cell extracts, while CB2 receptor bands are present in THP-1, HL-60 and microglial cell extracts.

Andis Klegeris, et al. Br J Pharmacol. 2003 June;139(4):775-786.
4.
Figure 7

Figure 7. From: Reduction of human monocytic cell neurotoxicity and cytokine secretion by ligands of the cannabinoid-type CB2 receptor.

The antineurotoxic effects of the specific CB2 receptor ligand JWH-015 on THP-1 cell toxicity towards SH-SY5Y neuronal cells are enhanced by REV 5901, when this specific inhibitor of 5-LOX is added at a suboptimal dose of 1 μM. Experiments were performed as described in Figure 5. The abscissa represents the concentration of JWH-015 in μM and the ordinate the percent survival of cells where 100% represents the survival in the presence of vehicle solution. Note that at 1 μM or less REV 5901 by itself had no significant effect in this assay (Klegeris & McGeer, 2002). Data (means±s.e.m.) are expressed as % live (a) or dead (b) cells, N=4, F and P-values indicate the level of significance, two-factor ANOVA.

Andis Klegeris, et al. Br J Pharmacol. 2003 June;139(4):775-786.
5.
Figure 6

Figure 6. From: Reduction of human monocytic cell neurotoxicity and cytokine secretion by ligands of the cannabinoid-type CB2 receptor.

The specific CB2 receptor ligand JWH-015 in a concentration-dependent manner inhibits neurotoxicity of human microglia derived from surgical specimens. Human microglial cells were pretreated with various concentrations of JWH-015 ( μM, shown on the abscissa) for 30 min before stimulation with LPS (0.5 μg ml−1) and IFN-γ (150 U ml−1). After 24 h incubation, the cell-free supernatants of microglial cells were transferred to the wells containing SH-SY5Y cells. The viability of SH-SY5Y cells was assessed after 72 h by the MTT assay (a) and by measuring the LDH activity in the supernatants (b). Data (means±s.e.m.) are expressed as % live (a) or dead (b) cells. The concentration-dependent effects of JWH-015 were assessed by randomized blocks design ANOVA. Data were obtained from four independent experiments, F and P-values obtained for both assays are presented in the figure. Note that human microglial cells were used at a five times lower concentration than THP-1 cells.

Andis Klegeris, et al. Br J Pharmacol. 2003 June;139(4):775-786.
6.
Figure 4

Figure 4. From: Reduction of human monocytic cell neurotoxicity and cytokine secretion by ligands of the cannabinoid-type CB2 receptor.

Toxicity of cannabinoids and synthetic ligands of cannabinoid receptors towards SH-SY5Y neuroblastoma cells. (a) SH-SY5Y cells were incubated with various concentrations of drugs (μM, shown on the ordinate) alone, and cell viability was assessed 72 h later by the MTT assay. (b) Toxic effect of Δ9-THC was abolished by the CB1 receptor antagonist SR141716A, but not by the CB2 receptor antagonist SR144528. Receptor antagonists (1μM) or their corresponding vehicle solutions were added 10 min before exposure of SH-SY5Y cells to 5 μM Δ9-THC. (c) Toxicity of anandamide was enhanced by inhibitors of the enzymatic hydrolysis of anandamide (PMSF and MAFP). Inhibitors or their corresponding vehicle solutions were added 10 min before exposure of SH-SY5Y cells to 50 μM anandamide. Data (means±s.e.m.) are expressed as percent control, where 100% (shown as a dashed line) is the value obtained in the presence of vehicle solution only. The dotted lines in (a) and (b) represent s.e.m. intervals. The number of independent experiments as well as P-values obtained for each of the drug treatments are presented on the figure. The concentration-dependent effects of various drugs in (a) were assessed by randomized blocks design ANOVA, while the effects of receptor antagonists in (b) and (c) were estimated by the paired Student's t-test and corrected for multiple comparisons by Holm's step-down method.

Andis Klegeris, et al. Br J Pharmacol. 2003 June;139(4):775-786.
7.
Figure 5

Figure 5. From: Reduction of human monocytic cell neurotoxicity and cytokine secretion by ligands of the cannabinoid-type CB2 receptor.

Decreased toxicity of THP-1 monocytic cell secretions towards SH-SY5Y cells by cannabinoids and synthetic ligands of cannabinoid receptors. THP-1 cells were pretreated with various concentrations of drugs (μM, shown on the ordinate) for 30 min before stimulation with LPS (0.5 μg ml−1) and IFN-γ (150 U ml−1). After 24 h incubation, the cell-free supernatants of THP-1 cells were transferred to the wells containing SH-SY5Y cells. The viability of SH-SY5Y cells was assessed after 72 h by the MTT assay (a) and by measuring the LDH activity in the supernatants (b). Data (means±s.e.m.) are expressed as % control, where 100% (shown as a dashed line) is the value obtained from supernatants of stimulated THP-1 cells in the presence of corresponding vehicle solution. The dash-dotted line represents the mean value obtained from supernatants of unstimulated THP-1 cells, while the dotted lines represent s.e.m. intervals. The number of independent experiments for each set of data is also shown. The concentration-dependent effects of various drugs were assessed by randomized blocks design ANOVA and P-values obtained for each of the drug treatments are presented on the figure. (c) The antineurotoxic effect of JWH-015 was abolished by the CB2 receptor antagonist SR144528 but not by the CB1 receptor antagonist SR141716A. Receptor antagonists (1 μM) or their corresponding vehicle solutions were added 10 min before exposure of THP-1 cells to 5 μM JWH-015. The number of independent experiments for each set of data is shown together with P-values that were obtained by the paired Student's t-test (versus THP-1 cells stimulated in the presence of vehicle solutions only) and corrected for multiple comparisons by Holm's step-down method.

Andis Klegeris, et al. Br J Pharmacol. 2003 June;139(4):775-786.

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