Table 1.
Primers used for realtime PCR.
Table 2.
Antibodies used for Western blots (WB) and Immunofluorescence (IF) labeling.
Figure 1.
Quantification of mitochondrial activity in hTM after 48 hours normalized to starting activity in controls. (A) Effects of ω-6 (16 µM) and ω-3 (50 µM) fatty acids compared to controls (Co). (B) Effects of H2O2 in controls, ω-6 and ω-3 fatty acids pre-treated hTM. (C) %-reduction of mitochondrial activity after H2O2 exposition. Values represent mean averages (m.a.) ± standard deviations (sd) of three independent experiments performed in triplicates of 5 different donors (n = 45); asterisks: p-values of statistical significances (**p≤0.01; ***p≤0.001).
Figure 2.
Quantification of proliferation rate in hTM after 48 hours normalized to starting activity in controls. (A) Effects of ω-6 (16 µM) and ω-3 (50 µM) fatty acids compared to controls (Co). (B) Effects of H2O2 in controls, ω-6 and ω-3 fatty acids pre-treated hTM. (C) %-reduction of BrdU-incorporation after H2O2 exposition. Values represent m.a. ± sd of three independent experiments performed in triplicates of 5 different donors (n = 45); asterisks: p-values of statistical significances (*p≤0.05; **p≤0.01; ***p≤0.001).
Figure 3.
Hsp27 and Hsp90 expression analysis.
(A) Quantification of realtime PCR expression analysis of Hsp27 and Hsp90 mRNAs in controls, ω-6 and ω-3 fatty acids pre-treated hTM normalized to controls. (B) Western blot detection of cellular Hsp27, Hsp90 and actin protein in controls, ω-6 and ω-3 fatty acids pre-treated hTM. (C) Plot of densitometric quantifications of Hsp27 and Hsp90 protein expression in controls, ω-6 and ω-3 fatty acids pre-treated hTM adjusted to actin expression and normalized to controls. Values represent m.a. ± sd of three independent experiments performed on cells of three different donors (n = 9); asterisks: p-values of statistical significances (*p≤0.05; **p≤0.01; ***p≤0.001).
Figure 4.
FN, PAI-1 and CTGF expression analysis.
Quantitative realtime PCR expression analysis of FN, PAI-1 and CTGF mRNAs in controls, ω-6 and ω-3 fatty acids pre-treated hTM normalized to controls (A) before and (B) after H2O2 exposition. (C) Western blot detection of cellular FN, PAI-1, CTGF, and actin in controls, ω-6 and ω-3 fatty acids pre-treated hTM. Plots of densitometric quantifications to deduce fold expressions of intracellular (D) FN, (E) PAI-1 and (F) CTGF, before and after H2O2 exposition. (G) ELISA quantification of FN medium contents normalized to controls. Values represent m.a. folds ± sd of three independent experiments performed on cells of three different donors (n = 9); asterisks: p-values of statistical significances (*p≤0.05; **p≤0.01; ***p≤0.001).
Figure 5.
Interleukin 1α, -6 and -8 expression analysis.
Quantitative realtime PCR expression analysis of interleukins (A) -1α, (B) -6 and (C) -8 mRNAs in controls, hTM pre-treated with ω-6 and ω-3 before and after H2O2 exposition. ELISA quantification of (D) IL-6 and (E) IL-8 medium contents. Values are normalized to untreated controls and represent m.a. folds ± sd of three independent experiments performed on cells of three different donors (n = 9); asterisks: p-values of statistical significances (*p≤0.05; **p≤0.01; ***p≤0.001).
Figure 6.
(A) Quantitative realtime PCR expression analysis of NFκB mRNA in controls, hTM pre-treated with ω-6 and ω-3 before and after H2O2 exposition. (B) Quantification of nuclear NFκB protein contents. Values are normalized to untreated controls and represent m.a. folds ± sd of three independent experiments performed on cells of three different donors (n = 9).
Figure 7.
Phalloidin labeling of the F-actin cytoskeleton in controls, ω-6 and ω-3 supplemented hTM (A) before and (B) after H2O2 exposition; scale bar: 100 µm.