Ongoing research at our Instituton includes work with 3D cell cultures based on two (so far) different extracellular matrix components. We have optimized protocols to prepare the 3D cultures from different cell lines (mainly cancer cell lines) and to visualize multicellular clusters and different intracellular organels in native 3D cultures by means of confocal fluorescence microscopy. We are studying cell morphogenesis, response of multicellular clusters to external stimuli, such as UV light illumination, photoinactivation mediated by photosensitizesrs as well as some of the biochemical activities, especially production of protoporhyrin IX from its precursors within the cells in extracellular matrix environment.
We are planning to extend the range of extracellular matrix components for preparation of 3D cell cultures to be able to study the effect of extracellular matrix on cellular processes in general. For this purpose we will also need to characterize our 3D cultures from biochemical and biophysical points of view. We have started with some preliminary experiments in this field, especially with visualization of collagen fibres surrounding cells in 3D cultures and image processing of the corresponding confocal images as well as with optimalization of protocols to prepare „tissue“ sections from the 3D cultures. We are planning to start with preliminary estimations of matrix stiffness in the near future.
Our future goal is to establish 3D co-cultures of different cell types, 3D cultures composed of different extracellular matrix components and modification of 3D cutlures using 3D microstructures prepared by 2-photon polymerisation technology to be able to model disease situations in vitro and thus to enable studies of cellular processes (mainly related to oncology) in clinically relevant conditions. This should help to evaluate experimental treatment approaches with potential benefits to patients.
The methods and techniques that we are commonly using include cell culturing, preparation of 3D cultures based on extracellular matrix components, confocal microscopy in combination with fluorescence staining, spectroscopic methods and visualisation and image processing of microscopic data from 3D cultures. These can be offered to other EuroCellNet participants at short notice.
Optimalization of methods for preparation of "tissue sections" from 3D cultures is in progress and hopefully could be offered to other EuroCellNet participants shortly.
We need help with establishing the method for estimation of extracellular matrix stiffness, preferably by AFM as one of the laboratories at our Institute is equiped with scanning probe microscope for biological samples.
Čunderlíková B Clinical significance of immunohistochemically detected extracellular matrix proteins and their spatial distribution in primary cancer. Crit Rev Oncol Hematol 105, 2016, 127-144.
Mateašík A, Trnka M, Kajo K, Vallová M, Čunderlíková B Cell-type dependent response to photodynamic treatment in 3D collagen cell cultures. J Photochem. Photobiol. Biology B, 2016, accepted.
Čunderlíková B Extracellular matrix in gene expression profiling of cancer. Transl Cancer Res 5, 2016, 44-48.
Čunderlíková B, Vasovič V, Sieber F, Furre T, Borgen E, Nesland JM, Peng Q. Hexaminolevulinate-mediated photodynamic purging of marrow grafts with murine breast carcinoma. Bone Marrow Transplant 46(8), 2011, 1118-27.
Čunderlíková B, Vasovič V, Sieber F, Furre T, Nesland JM, Peng Q. Hexaminolevulinate-mediated photodynamic purging of leukemia cells from BM. Bone Marrow Transplant 45(10), 2010, 1553-61.