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Functional assay of multidrug resistant cells using JC-1, a carbocyanine fluorescent probe
J M Kühnel1, J Y Perrot, A M Faussat
1BIOCIS, CNRS URA 1843, Faculté de Pharmacie, Châtenay Malabry, France.
Abstract:
Multidrug resistance (MDR) is characterized by a decrease in the efficiency of chemotherapeutic agents correlated with the expression and activity of a membrane protein: the permeability-glycoprotein (Pgp 170). Clinically, detection of MDR can be performed by functional tests based on the accumulation of fluorescent compounds such as rhodamine 123. With the aim of improving the sensitivity of such analysis, we have evaluated JC-1, a fluorescent lipophilic carbocyanine dye. Above a critical concentration, JC-1 aggregates in a 'liquid crystal' form. Aggregates display a specific red emission band centered at 597 nm whereas the monomers display a green emission band centered at 540 nm. JC-1 was avidly accumulated in sensitive K562 cells where it displayed both a green cytoplasmic and red mitochondrial fluorescence. In contrast, JC-1 was poorly accumulated in resistant K562 cells, which displayed only a slight green fluorescence. The level of JC-1 accumulation was correlated with the level of Pgp expression detected by MRK16 and UIC2 antibodies on a set of K562 subclones with increasing resistance levels. The specific fluorescence properties of JC-1 allow accurate discrimination between low-level resistant cells and sensitive cells. Chemosensitizers such as verapamil, cyclosporine A or S9788 restored JC-1 accumulation in resistant cells. The fluorescence properties of JC-1 could therefore be used for monitoring the effects of reversing agents.
Insights
This study introduces JC-1, a novel fluorescent dye, for detecting multidrug resistance (MDR) in cancer cells. JC-1 effectively distinguishes sensitive from resistant cells by its unique fluorescence properties, aiding in MDR detection.
Area of Science:
- Biochemistry
- Cell Biology
- Pharmacology
Background:
- Multidrug resistance (MDR) significantly reduces chemotherapy efficacy.
- The P-glycoprotein (Pgp 170) membrane protein is a key factor in MDR.
- Current MDR detection methods, like rhodamine 123 accumulation, have limitations in sensitivity.
Purpose of the Study:
- To evaluate JC-1, a novel fluorescent dye, for enhanced sensitivity in detecting MDR.
- To assess JC-1's ability to discriminate between sensitive and multidrug-resistant cancer cells.
- To explore JC-1's potential in monitoring the effects of MDR-reversing agents.
Main Methods:
- Utilized JC-1, a lipophilic carbocyanine dye, for fluorescence analysis in K562 cell lines.
- Investigated JC-1 accumulation and fluorescence emission (green monomers, red aggregates) in sensitive and resistant cells.
- Correlated JC-1 accumulation levels with Pgp 170 expression using MRK16 and UIC2 antibodies.
- Tested the effect of chemosensitizers (verapamil, cyclosporine A, S9788) on JC-1 accumulation in resistant cells.
Main Results:
- JC-1 accumulated significantly in sensitive K562 cells, showing green cytoplasmic and red mitochondrial fluorescence.
- Resistant K562 cells exhibited poor JC-1 accumulation with only slight green fluorescence.
- JC-1 accumulation levels directly correlated with Pgp 170 expression across resistant cell subclones.
- JC-1 fluorescence properties enabled accurate discrimination between low-level resistant and sensitive cells.
- Chemosensitizers restored JC-1 accumulation in resistant cells, indicating reversal of MDR.
Conclusions:
- JC-1 is a sensitive fluorescent probe for detecting and quantifying multidrug resistance in cancer cells.
- The distinct fluorescence of JC-1 monomers and aggregates allows for precise differentiation of resistant and sensitive cell populations.
- JC-1 facilitates monitoring the efficacy of MDR-reversing agents, offering potential for improved cancer therapy.