<em>In Vivo</em> Model for Testing Effect of Hypoxia on Tumor Metastasis

Abstract

Hypoxia has been implicated in the metastasis of Ewing sarcoma (ES) by clinical observations and in vitro data, yet direct evidence for its pro-metastatic effect is lacking and the exact mechanisms of its action are unclear. Here, we report an animal model that allows for direct testing of the effects of tumor hypoxia on ES dissemination and investigation into the underlying pathways involved. This approach combines two well-established experimental strategies, orthotopic xenografting of ES cells and femoral artery ligation (FAL), which induces hindlimb ischemia. Human ES cells were injected into the gastrocnemius muscles of SCID/beige mice and the primary tumors were allowed to grow to a size of 250 mm3. At this stage either the tumors were excised (control group) or the animals were subjected to FAL to create tumor hypoxia, followed by tumor excision 3 days later. The efficiency of FAL was confirmed by a significant increase in binding of hypoxyprobe-1 in the tumor tissue, severe tumor necrosis and complete inhibition of primary tumor growth. Importantly, despite these direct effects of ischemia, an enhanced dissemination of tumor cells from the hypoxic tumors was observed. This experimental strategy enables comparative analysis of the metastatic properties of primary tumors of the same size, yet significantly different levels of hypoxia. It also provides a new platform to further assess the mechanistic basis for the hypoxia-induced alterations that occur during metastatic tumor progression in vivo. In addition, while this model was established using ES cells, we anticipate that this experimental strategy can be used to test the effect of hypoxia in other sarcomas, as well as tumors orthotopically implanted in sites with a well-defined blood supply route.

Blocking OLFM4/HIF‐1α axis alleviates hypoxia‐induced invasion, epithelial–mesenchymal transition, and chemotherapy resistance in non‐small‐cell lung cancer

“…Hypoxia in solid tumors is a major negative prognostic factor for tumor progression because decreased availability of oxygen in tumors increases treatment resistance and enhances tumor progression and metastasis (Hong et al, 2016;Joseph, Harishankar, Pillai, & Devi, 2018;Ren et al, 2013). Adaptation to hypoxia has been shown to increase tumor cell invasiveness and metastatic potential, contributing to radio-or chemotherapeutic resistance (Klein et al, 2017;Shen et al, 2017).…”

The antitumor mechanisms of aerobic exercise: A review of recent preclinical studies

“… 40 Tumor cells have a propensity to metastasize under hypoxic conditions, and hypoxia downregulates the expression of adhesion molecules and increases tumor cell motility. 41 In addition, Rho is a redox sensitive small GTPase whose activation is associated with destabilization and increased permeability of the endothelial barrier during tumorigenesis. Tight junction proteins are major structural components that determine paracellular integrity of brain endothelium, which regulates endothelial cell permeability, and creates a barrier, through cell adhesion, that limits access to the cerebrospinal fluid environment.…”

“…The key steps in the initiation and growth of tumor metastasis depend on the ability of tumor cells to transverse the vascular membrane, enter the bloodstream, and penetrate endothelial cells at new sites 40 . Tumor cells have a propensity to metastasize under hypoxic conditions, and hypoxia downregulates the expression of adhesion molecules and increases tumor cell motility 41 . In addition, Rho is a redox sensitive small GTPase whose activation is associated with destabilization and increased permeability of the endothelial barrier during tumorigenesis.…”

Technical Feasibility and Physiological Relevance of Hypoxic Cell Culture Models

“…Certain tissue dyes such as Hypoxyprobe (pimonidazole hydrochloride) coupled with monoclonal antibodies are provided only with semi-quantitative assessments of hypoxia (4). Moreover, Hypoxyprobe is generally designed for use in patients and animal models to observe hypoxia of an explanted tissue (54,55), although sporadic use in vitro has also been reported (56). Nevertheless, there is another staining method that yields exact values of pO 2 , which is based on oxygenmediated quenching of the fluorescent signal that is inversely proportionate to pO 2 (57), the intensity of which is quantifiable by a microscope (58) or commercially available devices (59).…”