Fig. 6

Detoxification of ROS by adipose tissue extracellular fraction (AT-Ex) and plasma supplementation in normal human fibroblasts (NHF). a NHF were lysed after 24 and 72 h treatment for gene expression analysis of antioxidant enzymes by semiquantitative RT-PCR. β-actin expression was used to normalized cDNA concentration for each sample set. Means and SD are from six independent experiments. Statistical significance versus untreated control (Ctrl) is reported as *p < 0.05. b Gene expression data were confirmed at the protein level by Western blot analysis. Catalase (Cata) (c) and superoxide dismutase 2 (SOD2) (d) activity were measured using fluorescent and colorimetric assays, respectively. Samples were diluted 1:5 prior to assaying. Experiments were performed in duplicates. Histograms report data from single donors (n = 17 donors) and the mean value ± SD. Statistical significance versus untreated control is reported as *p < 0.05. e Mitochondrial membrane potential was measured by JC-1 staining and analyzed by flow cytometry. Increase in red fluorescence (monomer)/green fluorescence (aggregate) ratio demonstrated high mitochondrial membrane potential corresponding to more energized mitochondria. Means and SD are from six independent experiments. f Dot plot analysis of representative experiment shows a subpopulation of cells presenting depolarized mitochondria in plasma-treated NHF. Nac N-acetylcysteine