Electron spin resonance microscopic imaging of oxygen concentration in cancer spheroids

Hashem, Mada; Weiler-Sagie, Michal; Kuppusamy, Periannan; Neufeld, Gera; Neeman, Michal; Blank, Aharon

doi:10.1016/j.jmr.2015.04.012

Cited by 23 publications

(16 citation statements)

References 46 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…While our study shows oxygen as a percentage value in order to identify oxygenated 3D models to be used as an in vitro alternative to live fish tests, other studies report in terms of partial pressure. Oxygen partial pressure values of 50–60 mm Hg in the spheroid model have been reported in malignant cells using 3D oximetry methods [ 52 ]. Apart from the inherent differences between the two cell lines used (i.e.…”

Section: Resultsmentioning

confidence: 99%

Direct Measurements of Oxygen Gradients in Spheroid Culture System Using Electron Parametric Resonance Oximetry

et al. 2016

View full text Add to dashboard Cite

Advanced in vitro culture from tissues of different origin includes three-dimensional (3D) organoid micro structures that may mimic conditions in vivo. One example of simple 3D culture is spheroids; ball shaped structures typically used as liver and tumour models. Oxygen is critically important in physiological processes, but is difficult to quantify in 3D culture: and the question arises, how small does a spheroid have to be to have minimal micro-environment formation? This question is of particular importance in the growing field of 3D based models for toxicological assessment. Here, we describe a simple non-invasive approach modified for the quantitative measurement and subsequent evaluation of oxygen gradients in spheroids developed from a non-malignant fish cell line (i.e. RTG-2 cells) using Electron Paramagnetic Resonance (EPR) oximetry. Sonication of the paramagnetic probe Lithium phthalocyanine (LiPc) allows for incorporation of probe particulates into spheroid during its formation. Spectra signal strength after incorporation of probe into spheroid indicated that a volume of 20 μl of probe (stock solution: 0.10 mg/mL) is sufficient to provide a strong spectra across a range of spheroid sizes. The addition of non-toxic probes (that do not produce or consume oxygen) report on oxygen diffusion throughout the spheroid as a function of size. We provide evidence supporting the use of this model over a range of initial cell seeding densities and spheroid sizes with the production of oxygen distribution as a function of these parameters. In our spheroid model, lower cell seeding densities (∼2,500 cells/spheroid) and absolute size (118±32 μm) allow control of factors such as pre-existing stresses (e.g. ∼ 2% normoxic/hypoxic interface) for more accurate measurement of treatment response. The applied methodology provides an elegant, widely applicable approach to directly characterize spheroid (and other organoid) cultures in biomedical and toxicological research.

show abstract

Section: Resultsmentioning

confidence: 99%

Direct Measurements of Oxygen Gradients in Spheroid Culture System Using Electron Parametric Resonance Oximetry

et al. 2016

View full text Add to dashboard Cite

show abstract

“…Expression of CDK6 is regulated by different factors including hypoxia . A map of compact HCT‐116 tumour spheroids revealed spatial, locally restricted areas with reduced oxygen content . It is thus likely that similar locally restricted hypoxic conditions may account for reduced CDK6 expression in P114 tumour models.…”

Section: Discussionmentioning

confidence: 99%

In vitro study to assess the efficacy of CDK4/6 inhibitor Palbociclib (PD‐0332991) for treating canine mammary tumours

Schoos

Knab

Gabriel

et al. 2019

Vet Comparative Oncology

View full text Add to dashboard Cite

Therapy of canine mammary tumours (CMTs) with classical antitumour drugs is problematic, so better therapeutic options are needed. Palbociclib (PD-0332991) is an innovative and effective anticancer drug for the treatment of breast cancer in women. Palbociclib is an inhibitor of cyclin-dependent kinase 4 (CDK4) and CDK6, which are key regulators of the cell cycle machinery and thus cell proliferation. In the present in vitro study, we investigated whether Palbociclib also represents a candidate drug to combat CMT. For this purpose, the effect of Palbociclib was analysed in P114 and CF41 cells, two CMT cell lines with an endogenous CDK4/6 co-expression. Incubation of P114 and CF41 cells with Palbociclib resulted in a dose-and timedependent loss of phosphorylated retinoblastoma protein (pRb), a classical CDK4/6 substrate within the cell cycle machinery. Moreover, treatment of CMT cells with Palbociclib-induced cell cycle arrest affected cell viability, prevented colony formation and impaired cell migration activity. Palbociclib also inhibited the growth of P114 and CF41 cell spheroids. Immunohistochemical analysis of canine patient samples revealed a consistent expression of CDK6 in different canine mammary carcinoma types, but an individual and tumour-specific expression pattern of phosphorylated pRb independent of the tumour grade. Together, our findings let us suggest that Palbociclib has antitumour effects on CMT cells and that canine patients may represent potential candidates for treatment with this CDK4/6 inhibitor. K E Y W O R D S canine mammary tumour, CDK4/6, cell cycle, migration, Palbociclib

show abstract

“…Hypoxia, often combined with nutrient depletion and/or waste accumulation, collectively called "ischemia", is a key factor in determining cell survival and functioning in compact cell aggregates [39]. It is known that oxygen is consumed in vivo within about 200 microns distance, unless a blood supply is provided [40].…”

Section: Discussionmentioning

confidence: 99%

A heuristic computational model of basic cellular processes and oxygenation during spheroid-dependent biofabrication

et al. 2017

View full text Add to dashboard Cite

An emerging approach in biofabrication is the creation of 3D tissue constructs through scaffold-free, cell spheroid-only methods. The basic mechanism in this technology is spheroid fusion, which is driven by the minimization of energy, the same biophysical mechanism that governs spheroid formation. However, other factors such as oxygen and metabolite accessibility within spheroids impact on spheroid properties and their ability to form larger-scale structures. The goal of our work is to develop a simulation platform eventually capable of predicting the conditions that minimize metabolism-related cell loss within spheroids. To describe the behavior and dynamic properties of the cells in response to their neighbors and to transient nutrient concentration fields, we developed a hybrid discrete-continuous heuristic model, combining a cellular Potts-type approach with field equations applied to a randomly populated spheroid cross-section of prescribed cell-type constituency. This model allows for the description of: (i) cellular adhesiveness and motility; (ii) interactions with concentration fields, including diffusivity and oxygen consumption; and (iii) concentration-dependent, stochastic cell dynamics, driven by metabolite-dependent cell death. Our model readily captured the basic steps of spheroid-based biofabrication (as specifically dedicated to scaffold-free bioprinting), including intra-spheroid cell sorting (both in 2D and 3D implementations), spheroid defect closure, and inter-spheroid fusion. Moreover, we found that when hypoxia occurring at the core of the spheroid was set to trigger cell death, this was amplified upon spheroid fusion, but could be mitigated by external oxygen supplementation. In conclusion, optimization and further development of scaffold-free bioprinting techniques could benefit from our computational model which is able to simultaneously account for both cellular dynamics and metabolism in constructs obtained by scaffold-free biofabrication.

show abstract

Electron spin resonance microscopic imaging of oxygen concentration in cancer spheroids

Cited by 23 publications

References 46 publications

Direct Measurements of Oxygen Gradients in Spheroid Culture System Using Electron Parametric Resonance Oximetry

Direct Measurements of Oxygen Gradients in Spheroid Culture System Using Electron Parametric Resonance Oximetry

In vitro study to assess the efficacy of CDK4/6 inhibitor Palbociclib (PD‐0332991) for treating canine mammary tumours

A heuristic computational model of basic cellular processes and oxygenation during spheroid-dependent biofabrication

Contact Info

Product

Resources

About