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To detect apoptosis, the cells were stained with annexin?V and 7-amino actinomycin?D, and analyzed using flow cytometry. Reactive oxygen species, mitochondrial membrane depolarization and release of cytochrome?C into the cytosol were assessed using flow cytometry and enzyme-linked immunosorbent assay. Activity levels for the key apoptotic enzymes caspase-9, -8 and -3 were also determined. Genomic DNA damage was detected using single-cell gel electrophoresis. Results:? Apoptosis was significantly increased to 24.5 ��?5.7 at 24?h and 41.5 ��?8.9% at 48?h (p?Temsirolimus (CCI-779, NSC 683864) depolarization was collapsed. Cytochrome?C release was dramatically increased (0.12?��?0.02 vs. 0.02?��?0.01 at 24?h and 0.21?��?0.02 vs. 0.02?��?0.01 ng/mL at 48?h; p?http://www.selleckchem.com/products/pifithrin-alpha.html in human gingival fibroblasts. J Periodont Res 2012; 47: 701�C710. ? 2012 John Wiley & Sons A/S Background and Objective:? In our previous study, we found that flutamide [an androgen receptor (AR) antagonist] inhibited the up-regulation of collagen induced by interleukin (IL)-1�� and/or nifedipine in gingival fibroblasts. The present study attempted to verify the role of nitric Pfizer Licensed Compound Library oxide (NO) in the IL-1��/nifedipine-AR pathway in gingival overgrowth. Material and Methods:? Confluent gingival fibroblasts derived from healthy individuals (n?=?4) and those with dihydropyridine-induced gingival overgrowth (DIGO) (n?=?6) were stimulated for 48?h with IL-1�� (10?ng/mL), nifedipine (0.34?��m) or IL-1�� + nifedipine. Gene and protein expression were analyzed with real-time RT-PCR and western blot analyses, respectively. Meanwhile, Sircol dye-binding and the Griess reagent were, respectively, used to detect the concentrations of total soluble collagen and nitrite in the medium. Results:? IL-1�� and nifedipine simultaneously up-regulated the expression of the AR and type-I collagen ��1 [Col��1(I)] genes and the total collagen concentration in DIGO cells (p?
