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Subcellular Localization of Sulfonated Tetraphenyl Porphines in Colon Carcinoma Cells by Spectrally Resolved Imaging
Author(s) -
Malik Zvi,
Amit Ifat,
Rothmann Chana
Publication year - 1997
Publication title -
photochemistry and photobiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.818
H-Index - 131
eISSN - 1751-1097
pISSN - 0031-8655
DOI - 10.1111/j.1751-1097.1997.tb08576.x
Subject(s) - nucleus , fluorescence , subcellular localization , cytoplasm , vesicle , chemistry , biophysics , nucleolus , photochemistry , biochemistry , optics , biology , physics , membrane , microbiology and biotechnology
Subcellular localization of the dye, 5,10,15,20‐tetra(4‐sul‐fonatophenyl)porphine (TPPS 4 ) and the more hydrophobic dye, 5,10,15,20‐tetra(1‐sulfonatophenyl)porphine (TPPS 1 ), in murine colon carcinoma cells was studied by spectrally resolved imaging (SRI) combined with image processing techniques. Spectrally resolved imaging enabled the acquisition of multipixel fluorescence spectra (>10 4 ) from a single cell. Demarcation of specific localization sites and segregation of the irrelevant fluorescence were based on the pixel spectra and by operating the functions of spectral similarity mapping (SSM), principal component analysis (PCA) and spectral classification. The SRI revealed the fine details of the photochemical process that clarify some aspects of subcellular damage. The SRI depicted the differences between TPPS 4 and TPPS, with respect to their initial localization and their fate at the end of the photochemical effect. The dye TPPS 4 was localized initially in lysosomal vesicles, and upon irradiation fluorescence was seen in the nucleus as well as in vesicles. Some of the vesicles were closely related to the nucleus, as resolved by SSM, PCA and spectral classification. Additional light exposure stimulated relocalization of TPPS 4 into the nucleus as well as into the nucleolus, which was clearly depicted by SSM and PCA. Spectral classification showed a third, weak residual cytoplasmic array around the nucleus. The dye TPPS, concentrated in a Golgi‐like complex and was resolved in the nuclear envelope and in small vesicles: it was not redistributed into other compartments upon photosensitization. Serum supplementation to the incubation media of colon carcinoma cells treated with TPPS 4 or TPPS, did not change the localization patterns. Pixel spectra of the two dyes in the cells showed spectral shifts and expanded shoulders due to microenvironmental effects. Thus, the chemical nature of the sulfonated phenyl porphines, and not their interaction with serum proteins, was the main determinant of their binding to the lysosomes, nucleus, nucleolus, nuclear envelope or Golgi.