title: Estimation of PKCδ autophosphorylation in U87 MG glioma cells: combination of experimental, conceptual and numerical approaches.
abstract: Golgi apparatus (GA) is a center for lipid metabolism and the final target of ceramide pathway, which may result in apoptosis. In this work localization of highly hydrophobic hypericin is followed by time-resolved imaging of NBDC6(fluorescent ceramide) in U87 MG glioma cells. Decrease of NBDC6fluorescence lifetimes in cells indicates that hypericin can also follow this pathway. It is known that both, ceramide and hypericin can significantly influence protein kinase C (PKC) activity. Western blotting analysis shows increase of PKCδ autophosphorylation at Ser645 (p(S645)PKCδ) in glioma cells incubated with 500 nM hypericin and confocal-fluorescence microscopy distinguishes p(S645)PKCδ localization between GA related compartments and nucleus. Experimental and numerical methods are combined to study p(S645)PKCδ in U87 MG cell line. Image processing based on conceptual qualitative description is combined with numerical treatment via simple exponential saturation model which describes redistribution of p(S645)PKCδ between nucleus and GA related compartments after hypericin administration. These results suggest, that numerical methods can significantly improve quantification of biomacromolecules (p(S645)PKCδ) directly from the fluorescence images and such obtained outputs are complementary if not equal to typical used methods in biology.
authors.affiliation: Department of Biophysics, Faculty of Science, P.J. Safarik University (UPJS) in Kosice, Jesenna 5, 041 54, Kosice, Slovakia.
authors.forename: Matus
authors.initials: M
authors.lastname: Misuth
authors.affiliation: Department of Biophysics, Faculty of Science, P.J. Safarik University (UPJS) in Kosice, Jesenna 5, 041 54, Kosice, Slovakia.|Laboratory of Organometallic and Medicinal Chemistry, Institute of Chemical Sciences and Engineering, Swiss Federal Institute of Technology in Lausanne (EPFL), Station 6, Batiment de Chimie, CH-1015, Lausanne, Switzerland.
authors.forename: Jaroslava
authors.initials: J
authors.lastname: Joniova
authors.affiliation: Department of Biophysics, Faculty of Science, P.J. Safarik University (UPJS) in Kosice, Jesenna 5, 041 54, Kosice, Slovakia.
authors.forename: Dominik
authors.initials: D
authors.lastname: Belej
authors.affiliation: Department of Biophysics, Faculty of Science, P.J. Safarik University (UPJS) in Kosice, Jesenna 5, 041 54, Kosice, Slovakia.
authors.forename: Stanislav
authors.initials: S
authors.lastname: Hrivnak
authors.affiliation: Center for Interdisciplinary Biosciences, Faculty of Science, P.J. Safarik University (UPJS) in Kosice, Jesenna 5, 041 54, Kosice, Slovakia.
authors.forename: Denis
authors.initials: D
authors.lastname: Horvath
authors.affiliation: Center for Interdisciplinary Biosciences, Faculty of Science, P.J. Safarik University (UPJS) in Kosice, Jesenna 5, 041 54, Kosice, Slovakia.
authors.forename: Veronika
authors.initials: V
authors.lastname: Huntosova
mesh_terms: D000873:Anthracenes; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000201:enzymology / Q000473:pathology; D002518:Ceramides / Q000378:metabolism; D003198:Computer Simulation; D005910:Glioma / Q000201:enzymology* / Q000473:pathology; D006801:Humans; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D061848:Optical Imaging; D010569:Perylene / Q000031:analogs & derivatives / Q000378:metabolism; D010766:Phosphorylation; D051745:Protein Kinase C-delta / Q000378:metabolism*; D015398:Signal Transduction; D013057:Spectrum Analysis; D013997:Time Factors
chemical_list: D000873:Anthracenes; D002518:Ceramides; D010569:Perylene; C004965:hypericin; D051745:Protein Kinase C-delta
keywords: Golgi apparatus; ceramide; hypericin; numerical analysis; protein kinase C delta