merocyanine-dye and Colonic-Neoplasms

The colon cancer tissues from DMH treated rats exhibited higher membrane potential, fluidity and changed lipid order as examined by Merocyanine 540 and 1,6-diphenyl-1,3,5-hexatriene, respectively. A transition from gel to liquid crystalline state was observed by Laurdan fluorescence and also reduced fluorescence quenching of NBD-PE as contributed in the decreased membrane lipid phase separation. With piroxicam, a traditional NSAID and c-phycocyanin, a biliprotein from Spirulina platensis, these effects were normalized. An augmented intracellular Ca(+2) had contributed to the drug mediated apoptosis which is supported by an elevated calpain-9 expression. Histopathologically, a large pool of secreted acid/neutral mucopolysaccrides as well as the presence of blood vessels and dysplastic crypts signifies invasive mucinous adenocarcinoma while both the drugs reduced these neoplastic alterations. Wnt/β-catenin pathway was also found to be up-regulated which served as a crucial indicator for cancer cell growth. A concomitant down regulation of PPARγ was noted in DMH treatment which is associated with tumor progression. The expression of PPARα and δ, the other two isoforms of PPAR family was also modulated. We conclude that piroxicam and c-phycocyanin exert their anti-neoplastic effects via regulating membrane properties, raising calpain-9 and PPARγ expression while suppressing Wnt/β-catenin signaling in experimental colon carcinogenesis.

Topics: 2-Naphthylamine; Animals; Apoptosis; Calcium; Calpain; Carcinogenesis; Colonic Neoplasms; Fluorescence Polarization; Fura-2; Intracellular Space; Laurates; Ligands; Male; Membrane Fluidity; Membrane Potentials; Phase Transition; Phosphatidylethanolamines; Phycocyanin; Piroxicam; PPAR gamma; Pyrimidinones; Rats, Sprague-Dawley; Up-Regulation; Wnt Signaling Pathway

It has been suggested that selective uptake of photosensitizers is due to significantly lower pH of the interstitial fluid in tumors compared to normal tissue. Therefore, the cellular uptake of merocyanine 540 (MC 540) was examined at two pH values: 6.8+/-0.1 and 7.4+/-0.1. There was no difference in spectral properties (absorption and fluorescence maxima positions, fluorescence intensity) of the drug in the presence of increasing amounts of either human blood plasma or FCS (0-2%) at the two pH values investigated. Nevertheless, significantly higher amounts of the drug were taken up by WiDr cells at pH 6.8+/-0.1, both in the presence of 10% FCS and in the absence of FCS. The absorption spectra of MC 540 in the presence of egg phosphatidylcholine (PC) liposomes turned out to be NaCl concentration-dependent (0.00-0.30 mol l(-1)). Membrane fluidity, as measured by fluorescence anisotropy of diphenylhexatriene (DPH), was unchanged within the experimental error in the NaCl concentration range 0.01-0.30 mol l(-1). The spectral changes indicated an enhancement of the incorporation of MC 540 into lipid membranes with increasing ionic strength. Such a salt concentration dependence suggests a possible involvement of the surface potential in the interaction of MC 540 with lipid membranes. The results might provide an explanation of the pH dependency of the cellular uptake of MC 540 observed in this study.

Topics: Adenocarcinoma; Blood Proteins; Cell Membrane Structures; Colonic Neoplasms; Culture Media; Humans; Hydrogen-Ion Concentration; Ions; Lipid Bilayers; Liposomes; Photosensitizing Agents; Pyrimidinones; Spectrometry, Fluorescence; Tumor Cells, Cultured

The relationship between tumor cell differentiation and photosensitizer accumulation used in PDT is poorly defined. In the present work, specific cell differentiation of colon carcinoma CT26 cells induced by sodium butyrate was manifested by morphological changes, proliferation and protein expression and was correlated with the accumulation of endogenous and exogenous photosensitizes. Reduced accumulation of the endogenous protoporphyrin IX and the exogenous hypericin and MC540 was detected in differentiated cells. In contrast, a differentiation-dependent increase was measured with TPPS4, TMPyP, the pheophorbides (C5, C6, C12), HypS4 and helianthrone. In conclusion, PpIX, Hypericin and MC540 show specific binding and accumulation in poorly differentiated tumors, giving these tumors tissue-specific advantage in photo-diagnostic PDT applications.

Topics: Animals; Anthracenes; Butyrates; Carcinoma; Cell Cycle; Cell Differentiation; Colonic Neoplasms; Mice; Perylene; Photosensitizing Agents; Protoporphyrins; Pyrimidinones; Tumor Cells, Cultured

We studied the staining pattern of merocyanine 540 (MC540) by spectral imaging of murine CT26 and human HT29 colon carcinoma cells incubated with the dye MC540. This dye, usually considered a potential membrane probe, localized mainly in the cytoplasmic vesicles of the colon carcinoma cells. However, in cells incubated in an environment similar to that of a tumor (pH 6.7), high fluorescence was detected in the nuclear membrane and nucleoli. Under these acidic conditions, resembling the Krebs effect, a population of CT26 cells displayed fluorescent plasma membranes. In differentiating cells, exhibiting cell cycle arrest at G(1)-phase and an elevated level of alkaline phosphatase, MC540 fluorescence was confined to cytoplasmic vesicles and was not detected in the plasma membrane or in the nucleoli. Cell sorting analysis of both cell types at pH 5.0 revealed higher fluorescence intensity in proliferating cells compared to differentiating cells. The fluorescence intensity of MC540-stained cells reached a maximum at pH 5.0, although the fluorescence of MC540 dye was maximal at pH 7.2. This phenomenon may result from increased binding of MC540 monomers to the cells because disaggregation of the dye with Triton X-100 produced similar results. We conclude that nucleolar localization of MC540 and an elevated fluorescence intensity can be used as indicators for proliferating cells in the characteristically acidic tumor environment. (J Histochem Cytochem 49:147-153, 2001)

Topics: Alkaline Phosphatase; Animals; Butyrates; Cell Differentiation; Cell Division; Colonic Neoplasms; Flow Cytometry; Fluorescent Dyes; Humans; Hydrogen-Ion Concentration; Mice; Pyrimidinones; Spectrometry, Fluorescence; Subcellular Fractions; Tumor Cells, Cultured