Fig. 4
Post-glycated matrices support fibroblast viability for long periods. Primary human lung fibroblasts cultured within different glycated collagen DMEM matrices over a period of 21 days. a. Fluorescence determination using alamarBlue assay. b. LIVE/DEAD viability/cytotoxicity stained cells observed using confocal reflection microscopy. The viable cells are stained green, and the non-viable cells are stained red. a and b. Cell death occurred under all conditions at the highest ribose (R) concentration (240 mM), independently of the use of serum (FBS). The use of FBS increased the proliferation rate under the rest of the conditions (p < 0.01). a1 and b1 (0 % FBS). No cell proliferation was observed, but cells were alive during the 21 days. a2 and b2 (1 % FBS). A gradual increase in cell proliferation was observed under all conditions except R 240 mM, but the rates for R 5 mM (p < 0.05) and the control matrices (p < 0.01) were significantly different only between the 7th and 21st days. a3 and b3 (10 % FBS). An improved proliferation rate was observed with the use of 10 % of serum, which was significantly for the control (non-glycated) only between the 7th and 14th days (p < 0.05). Gel contraction was observed in ≤15 mM R conditions between the 14th and the 21st day. No gel contraction was observed at 30 mM R (explanation in discussion). It was not possible to determine the fluorescence in the controls and in matrices glycated using 5 mM R at the 21st day because of gel contraction and cells growing on the bottom of the well (a3). The scale bars correspond to 200 μm. The experiments were repeated three times, with similar results obtained