Estimation of oxygen and glucose metabolism and glucose diffusivity.

(A) Simulated oxygen concentrations as a function of distance from the centre of the spheroid during growth under standard growth conditions of replacing 50% (100 μL) of the culture medium every 2nd day based on experimentally derived oxygen consumption rate and literature medium and tissue diffusion coefficients (see Table 1 and supplementary material). (B) Glucose consumption by HCT116 monolayers (105 cells in 0.2 mL/well) in glucose-free medium with no FCS and a range of D-glucose concentrations were measured by serially sampling 5 μL medium at each time point. Values are means ± SD for 4 replicates. D-glucose metabolism rate parameters (Vmax and Km) in HCT116 cells were then calculated using the MABM (lines) using parameters that minimize the sum of squared errors between observed and simulated glucose concentrations for the full data set simultaneously. (C) Correlation between observed and MABM predicted glucose concentration data in B (R2 = 0.977) using the optimized glucose metabolism parameters. (D) Schematic figure of apparatus for the measurement of glucose and drug diffusion and metabolism through MCLs grown on a porous support membrane. After drug is added to the donor compartment, samples were collected from the donor and receiver compartments at intervals for measurement of concentrations of glucose or drug and its metabolite(s). (E) Representative diffusion data showing transport of 3H-L-glucose or D-glucose through HCT116 MCL (grown for 3 days, ca 100 μm thickness). Concentrations are normalised to the initial concentration measured in the donor compartment. Lines are model fits (minimization of the sum of squared errors between observed and calculated concentrations) with fitted parameters Vmax for D-glucose, after fixing the glucose coefficient at its mean value determined for L-glucose curves. (F) Simulated D-glucose concentration as a function of distance from the centre of the spheroid during growth under standard growth conditions of replacing 50% (100 μL) of the culture medium every 2nd day.