Figure 7
Effects of various compounds on the secretion of TNF-α, in LPS-stimulated (pre-treatment) peritoneal macrophages of 8-week-old C57BL/6 versus BALB/c female mice. Thioglycolate-elicited peritoneal macrophages were prepared from 8-week-old C57BL/6 and BALB/c (5 of each) female mice as described previously [6]. The macrophages of each mouse were adhered to the bottom of 12 well plates (1 × 107 cells/well in 1 mL media) for 4 h, the cells were washed with media three times. The cells were cultured overnight in the fresh media (500 μL) after the final wash. The cells were treated with 100 μL dissolved in 0.2% DMSO of dexamethasone, 10 μM; mevinolin, 20 μM; δ-tocotrienol, 10 μM; α-tocopherol, 100 μM; riboflavin, 40 μM; or quercetin-HCL, 40 μM for I h, followed by stimulation with LPS (10 n g/mL) of each treatment. The assay mixtures were incubated at room temperature for 4 h, and the assay mixtures were centrifuged at 2,000 rpm for 20 min. The cells were then harvested, and the total cellular RNA was extracted from each pellet with RNeasy mini kit according to manufacturer's instructions [7, 17]. The total secretion of TNF-α of each inhibitor was estimated in the supernatants by radio-immunoassay (ELISA) kit according to the manufacture directions [7, 17]. The cells viability were very good (> 95%) in all the treatments [4, 7]. Data are means ± SD, n = 5 per treatment, and triplicate analyses of each sample. The treatments 1-7 correspond to: 1. Control (macrophages + LPS + 0.2% DMSO); 2. dexamethasone; 3. mevinolin; 4. α-tocopherol; 5. δ-tocotrienol; 6. riboflavin; 7. Quercetin-HCL. Values in a column with a different superscript symbols are significantly different at P< 0.05, A = values, B = Percentages.