Invasion Chemotaxis

Key words Invasion, Chemotaxis, Tumor microenvironment, In vivo, Chemoattractant... 

Key words Migration, Haptotaxis, Chemotaxis, Invasion, Motility, Matrix protein, 3-dimensional... 

Amnion mounting. Use a 24-well two-compartment chemotaxis chamber apparatus, with 1.6 cm well diameter. Stretch and clamp the denuded BM, either labeled or nonlabeled, within the holders, thus dividing the upper and lower compartments (Fig. 4.3). For invasion studies, sandwich a Millipore filter (8 /xm pore size) in direct contact with the amnion stromal surface. 

Although at the present time, relatively little is known regarding the signalling pathways activated by chemokines in cancer cells, preliminary data show that CXCR4 and CCR7 are capable of activating a number of different intracellular events such as chemotaxis, invasion and adhesion, all of which are properties that correlate with metastatic behaviour of tumour cells [40, 95]. 

Reiland, J., L. T. Furcht and J. B. McCarthy (1999). CXC-chemokines stimulate invasion and chemotaxis in prostate carcinoma cells through the CXCR2 receptor. Prostate 41(2) 78-88. 

The Boyden chamber is a simple apparatus used to test for chemotaxis, especially of leukocytes. It can also be used to assess tumor cell transmigration across an endo-thehal monolayer in vitro. It consists of two compartments separated by a MiUi-pore filter (3-8 pm pore size). A chemotac-tic factor is placed in one compartment, and a gradient develops across the thickness of the filter (ca. 150 pm). Cell movement into the filter is measured after an incubation period less than the time taken for the gradient to decay. Cell motility can be measured in Boyden chambers containing filters precoated with different materials, for example fibronectin or fibronectin fragments. The method, when apphed to malignant and non-mahgnant cell hnes, shows that the variable invasive potentials of these cells correlate with their abihty to disrupt the endothelial cell monolayer. 

Zebrafish have emerged as a powerfiil model organism to study neutrophil chemotaxis and inflammation in vivo. Studies of neutrophil chemotaxis in animal models have previously been hampered both by the limited number of specimens available for analysis and by the need for invasive procedures to perform intravital microscopy. Due to the transparency and cell permeability of zebrafish embryos these limitations are circumvented, and the zebrafish system is amenable to both live time-lapse imaging of neutrophil chemotaxis and for screening of the effects of chemical compounds on the inflammatory response in vivo. Here, we describe methods to analyze neutrophil-directed migration toward wounds using both fixed embryos by myeloperoxidase activity assay, and live embryos by time-lapse microscopy. Further, methods are described for the evaluation of the effects of chemical compounds on neutrophil motility and the innate immune responses in zebrafish embryos. 

Asymmetric localization of intracellular proteins and signals directs movement dimng axon guidance, endothelial cell invasion, and immune cell migration. In these processes, cell movement is guided by external chemical cues in a process known as chemotaxis. In particular, leukocyte migration in the innate immune system has been studied in the human neutrophil-like cell line (HL-60). Here, we describe the maintenance and transfection of HL-60 cells and explain how to analyze their behavior with two standard chemotactic assays. Finally, we demonstrate how to fix and stain the actin cytoskeleton of polarized cells for fluorescent microscopy imaging. 

To determine the optimal concentration for collecting the invasive population, it is often necessary to perform dose response experiments and try a range of chemoattractant amounts. We have found that for EGF, the optimal concentration in the in vivo invasion assay is 25 nM while for the in vitro Boyden chamber chemotaxis it is 5 nM. The difference can be explained by different diffusion properties of the gradient emanating from a tip of a needle into live tissue vs. EGF in a buffer being directly accessible to migrating cells across pores in a filter. By the same token, the optimal... 

S. (2003) Gene expression analysis on small numbers of invasive cells collected by chemotaxis from primary mammary tumors of the mouse. BMCBiotechnol. 3, 13. 

Vene R, Benelli R, Noonan DM, Albini A HIV-Tat dependent chemotaxis and invasion, key aspects of tat mediated pathogenesis. Clin Exp Metastasis 2000 18 533-538. 

B. Mosadegh, et al., A paper-based invasion assay assessing chemotaxis of cancer cells in gradients of oxygen. Biomaterials 52 (2015) 262-271. 

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