Rapid uptake of glucose and lactate, and not hypoxia, induces apoptosis in three-dimensional tumor tissue culture

Abstract

The spatial arrangement of cellular metabolism in tumor tissue critically affects the treatment of cancer. However, little is known about how diffusion and cellular uptake relate to intracellular metabolism and cell death in three-dimensions. To quantify these mechanisms, fluorescent microscopy and multicellular tumor cylindroids were used to measure pH and oxygen profiles, and quantify the distribution of viable, apoptotic and necrotic cells. Spheroid dissociation, enzymatic analysis, and mass spectrometry were used to measure concentration profiles of glucose, lactate and glutamine. A mathematical model was used to integrate these measurements and calculate metabolic rate parameters. It was found that large cylindroids, >500μm in diameter, contained apoptotic and necrotic cells, whereas small cylindroids contained apoptotic but not necrotic cells. The center of cylindroids was found to be acidic but not hypoxic. From the edge to the center, concentrations of glucose, lactate and glutamine decreased rapidly. Throughout the cell masses lactate was consumed and not produced. These measurements indicate that apoptosis was the primary mechanism of cell death; acidity was not caused by lactic acid; and cell death was caused by depletion of carbon sources and not hypoxia. The mathematical model showed that the transporter enzymes for glucose and lactate were not saturated; oxygen uptake was limited by intracellular metabolism; and oxygen uptake was not limited by membrane-transport or diffusion. Unsaturated transmembrane uptake may be the cause of both proliferative and apoptotic regimes in cancer. These results suggest that transporter enzymes are excellent targets for treating well oxygenated tumors.

Engineered bacteria detect spatial profiles in glucose concentration within solid tumor cell masses

“…A and B). This profile mimics previous measurements of glucose concentration in tumor cylindroids, in which concentration also dropped exponentially with depth into tissue (Kasinskas et al, ).…”

“…The average glucose concentration profile (red line) was used to estimate the consumption rate of glucose by the cancer cells. ( C ) Tumor tissue viability as a function of tissue depth as measured by caspase‐3 activity (data from Kasinskas et al, ). The average value was fit to continuous function for mathematical analysis of tumor treatment.…”

Comparison of cancer cells cultured in 2D vs 3D reveals differences in AKT/mTOR/S6-kinase signaling and drug response

“…The inner regions of spheroids experience decreased nutrient availability (such as amino acids), which translates into diminished nutrient signaling and is mandatory for mTOR activation (Jewell et al, 2013). Indeed, intracellular amino acid concentrations rapidly drop along a gradient to the insides of spheroids (Kasinskas et al, 2014). This might explain the drastic decrease of RPS6 phosphorylation -from high levels in the outer two-cell-layer-thick zone to low levels in the deeper areas.…”

Trg-deficient Salmonella colonize quiescent tumor regions by exclusively penetrating or proliferating

“…Without Trg, Salmonella were not attracted to extracellular glucose, which is at high concentrations in tumor tissue near blood vessels [34]. Colony localization of trg − Salmonella is dominated by attraction to other small molecules [20].…”