Comparative pO2 measurements in cell spheroids cultured with different techniques

Acker, H.; Carlsson, Jörgen; Mueller-Klieser, W.; Sutherland, RJ

doi:10.1038/bjc.1987.197

Cited by 48 publications

(30 citation statements)

References 6 publications

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“…For glucose D ~ 1×10 −7 cm 2 /sec [37] and uptake rate may be as large as 1×10 −3 sec −1 [38], while for oxygen the corresponding values are ~ 1×10 −5 cm 2 /sec [39] and 1×10 −1 sec −1 [40]. Diffusion penetration lengths thus calculated ( L ≈ 100 µm) are consistent with our experimental observations (IHC) and with previous (murine mammary tumor EMT6/Ro spheroid) observations showing glucose concentration decreasing by 65% [41] and oxygen decreasing by 90% [42] across the tumor viable region.…”

Section: Methodssupporting

confidence: 89%

Prediction of Drug Response in Breast Cancer Using Integrative Experimental/Computational Modeling

Edgerton²,

et al. 2009

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Nearly 30% of women with early-stage breast cancer develop recurrent disease attributed to resistance to systemic therapy. Prevailing models of chemotherapy failure describe three resistant phenotypes: cells with alterations in transmembrane drug transport, increased detoxification and repair pathways, and alterations leading to failure of apoptosis. Proliferative activity correlates with tumor sensitivity. Cell-cycle status, controlling proliferation, depends on local concentration of oxygen and nutrients. Although physiologic resistance due to diffusion gradients of these substances and drugs is a recognized phenomenon, it has been difficult to quantify its role with any accuracy that can be exploited clinically. We implement a mathematical model of tumor drug response that hypothesizes specific functional relationships linking tumor growth and regression to the underlying phenotype. The model incorporates the effects of local drug, oxygen, and nutrient concentrations within the three-dimensional tumor volume, and includes the experimentally observed resistant phenotypes of individual cells. We conclude that this integrative method, tightly coupling computational modeling with biological data, enhances the value of knowledge gained from current pharmacokinetic measurements, and, further, that such an approach could predict resistance based on specific tumor properties and thus improve treatment outcome. [Cancer Res 2009;69(10):4484-92]

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Section: Methodssupporting

confidence: 89%

Prediction of Drug Response in Breast Cancer Using Integrative Experimental/Computational Modeling

Edgerton²,

et al. 2009

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“…Supporting this prediction, we have shown that the primary oxygen-responsive transcription factor, HIF-1α, does not affect metabolism in the absence of glucose 18 . In spheroids, the presence of central hypoxia depends on cell type 19 . Spheroids have been formed that have both hypoxic 20 and well-oxygenated 21 cores.…”

Section: Introductionmentioning

confidence: 99%

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

2014

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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.

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“…4C, Lower and Fig. 4D, red line), coinciding with the diffusion limit of oxygen in spheroid masses (40). RT-PCR analysis demonstrated that HIF-2α mRNA expression was reduced in DNMT3a-expressing spheroids (Fig.…”

Section: Mrna (mentioning

confidence: 99%

DNMT3a epigenetic program regulates the HIF-2α oxygen-sensing pathway and the cellular response to hypoxia

Lachance

Uniacke

Audas

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Epigenetic regulation of gene expression by DNA methylation plays a central role in the maintenance of cellular homeostasis. Here we present evidence implicating the DNA methylation program in the regulation of hypoxia-inducible factor (HIF) oxygensensing machinery and hypoxic cell metabolism. We show that DNA methyltransferase 3a (DNMT3a) methylates and silences the HIF-2α gene (EPAS1) in differentiated cells. Epigenetic silencing of EPAS1 prevents activation of the HIF-2α gene program associated with hypoxic cell growth, thereby limiting the proliferative capacity of adult cells under low oxygen tension. Naturally occurring defects in DNMT3a, observed in primary tumors and malignant cells, cause the unscheduled activation of EPAS1 in early dysplastic foci. This enables incipient cancer cells to exploit the HIF-2α pathway in the hypoxic tumor microenvironment necessary for the formation of cellular masses larger than the oxygen diffusion limit. Reintroduction of DNMT3a in DNMT3a-defective cells restores EPAS1 epigenetic silencing, prevents hypoxic cell growth, and suppresses tumorigenesis. These data support a tumor-suppressive role for DNMT3a as an epigenetic regulator of the HIF-2α oxygensensing pathway and the cellular response to hypoxia.

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Comparative pO2 measurements in cell spheroids cultured with different techniques

Cited by 48 publications

References 6 publications

Prediction of Drug Response in Breast Cancer Using Integrative Experimental/Computational Modeling

Prediction of Drug Response in Breast Cancer Using Integrative Experimental/Computational Modeling

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

DNMT3a epigenetic program regulates the HIF-2α oxygen-sensing pathway and the cellular response to hypoxia

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