Influence of Cell Detachment on the Respiration Rate of Tumor and Endothelial Cells

Abstract

Cell detachment is a procedure routinely performed in cell culture and a necessary step in many biochemical assays including the determination of oxygen consumption rates (OCR) in vitro. In vivo, cell detachment has been shown to exert profound metabolic influences notably in cancer but also in other pathologies, such as retinal detachment for example. In the present study, we developed and validated a new technique combining electron paramagnetic resonance (EPR) oximetry and the use of cytodex 1 and collagen-coated cytodex 3 dextran microbeads, which allowed the unprecedented comparison of the OCR of adherent and detached cells with high sensitivity. Hence, we demonstrated that both B16F10 melanoma cells and human umbilical vein endothelial cells (HUVEC) experience strong OCR decrease upon trypsin or collagenase treatments. The reduction of cell oxygen consumption was more pronounced with a trypsin compared to a collagenase treatment. Cells remaining in suspension also encounter a marked intracellular ATP depletion and an increase in the lactate production/glucose uptake ratio. These findings highlight the important influence exerted by cell adhesion/detachment on cell respiration, which can be probed with the unprecedented experimental assay that was developed and validated in this study.

Cancer metabolism in space and time: Beyond the Warburg effect

“…In ad di tion to apop to sis, var i ous other bi o log i cal processes can lead to cell de struc tion. Anoikis is re lated to apop to sis as it is a cas pase de pen dent cell death mech a nism trig gered by the de tach ment of cells from the ex tra cel lu lar ma trix [9,110,111]. Necro sis, as op posed to the highly reg u lated apop to sis, is an ac ci den tal death oc cur ring spon ta neously fol low ing ex ter nal stresses that block nor mal phys i o log i cal processes.…”

“…In ter est ingly, cell de tach ment from the ex tra cel lu lar ma trix and neigh bor ing cells is per se able to in duce early meta bolic changes, in clud ing di ver sion of glu cose from gly col y sis and the TCA cy cle to the PPP [9], and de creases the oxy gen con sump tion rate [111]. The PPP is an an a bolic path way that pro duces NADPH, a co fac tor nec es sary for glu tathione re cy cling and ROS detox i fi ca tion.…”

Metabolic Markers and HSP60 in Chemonaive Serous Solid Ovarian Cancer Versus Ascites

“…10,11 In vitro studies of melanoma and endothelial cell lines have demonstrated cell detachment to result in decreased respiration and glucose consumption, implying metabolic reprogramming toward a less oxidative phenotype. 12,13 Despite a lower glucose uptake, these detached cells maintain lactate production, indicating use of alternative nutrients, for example, glutamine as energy substrates. 13 Altogether, these findings support the idea of malignant cell metabolic plasticity.…”

“…12,13 Despite a lower glucose uptake, these detached cells maintain lactate production, indicating use of alternative nutrients, for example, glutamine as energy substrates. 13 Altogether, these findings support the idea of malignant cell metabolic plasticity.…”

In Situ EPR Detection of Reactive Oxygen Species in Adherent Cells Using Polylysine-Coated Glass Plate

“…Some reports have shown that the amount of superoxide generated from leukocytes in suspension is proportional to a cell density, whereas it is inversely related to the cell density in adherent cells due to the mutual contact-induced suppression of the radical generation [22]. Another study proved that detached cells consume significantly less oxygen than adherent cells in both murine melanoma and human umbilical vein endothelial cells, indicating a considerable effect of the adhesion/ detachment process on the cell respiration rate [23]. It is therefore necessary to develop an EPR microscopy that enables us to accurately examine the redox state of a cell in the adhesion state, i.e., in situ, in combination with a conventional optical microscope.…”