Table 2.

Effect of HCT1026 on ageing-induced increased reactive microglial cell markers and survival assays

Treatments	DCFH-DA (% of control)	Nitrite (% of control)	MTT (% of control)	Caspase3 (% of control)
Plus PBS	220 ± 22	205 ± 18	95 ± 10	105 ± 12
Plus HCT1026
5 μm	170 ± 24*	156 ± 15*	100 ± 9	98 ± 10
10 μm	105 ± 14*	110 ± 10*	102 ± 12	100 ± 14
25 μm	90 ± 12*	87 ± 11*	95 ± 8	110 ± 8

Microglial (IBA1⁺) cells acutely isolated from young (control) and aging mouse brain were cultured for 24–48 h in the presence of the NO-donating NSAID, HCT1026 [2-fluoro-α-methyl(1,1'-biphenyl)-4-acetic-4-(nitrooxy)butyl ester] or phosphate buffered saline (PBS), as described. Experiments were performed with the freshly prepared IBA1⁺ cells cultured in the presence of increasing doses of HCT1026 (5–25 μm) applied after plating. The cells were processed 24–48 h after plating. Part of the cultures were processed for intracellular ROS using the redox membrane-permeant probe 2',7'-dichlorofluorescein diacetate (DCFH-DA, 50 μm, added for 1 h at 37°C), and cells viewed under the confocal microscope (Gennuso et al. 2004). Measurement of iNOS-derived NO was carried out in cell-free supernatant using Griess reagent (Marchetti et al. 2002; Morale et al. 2004; L'Episcopo et al. 2011c, 2012). Cell survival was determined by the MTT and Caspase3 assays. Results are expressed as percent changes over control (young microglial treated with PBS = 100). *p < 0.05 versus PBS.