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Fluorescence Polarization of Methylene Blue as a Quantitative Marker of Breast Cancer at the Cellular Level.

ABSTRACT: A quantitative technique to detect cancer in single cells could transform cancer diagnosis. Current cancer diagnosis utilizes histopathology, which requires tissue acquisition, extensive processing and, in most cases, relies on the qualitative morphological analysis of tissues and cells. Molecular biomarkers are only available for a few specific tumor subtypes. We discovered that the fluorescence polarization (Fpol) of Methylene Blue (MB) is significantly higher in cancer than in normal human breast tissues and cells. We confirmed that fluorescence polarization imaging did not affect the viability of the cells and yielded highly significant differences between cancer and normal cells using MB concentrations as low as 0.05 and 0.01 mg/ml. To explain this phenomenon we examined intracellular localization of MB and its fluorescence lifetime. We determined that higher fluorescence polarization of MB occurs due to its increased accumulation in mitochondria of cancer cells, as well as shorter fluorescence lifetime in cancer relative to normal cells. As quantitative MB Fpol imaging can be performed in vivo and in real time, it holds the potential to provide an accurate quantitative marker of cancer at the cellular level.

SUBMITTER: Yaroslavsky AN

PROVIDER: S-EPMC6353996 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:A promising alternative to antibiotics for treatment of Staphylococcus aureus infections is photodynamic inactivation (PDI), which employs a photosensitizer (PS) that produces cytotoxic reactive oxygen species (ROS) when exposed to molecular oxygen and antimicrobial blue light in the spectrum of 400-470 nm. Although the precise mechanistic basis of PDI has not been defined, the formation of ROS and free radicals that oxidize a number of cellular targets, including membrane lipids, damage to proteins and nucleic acids result in inactivation of essential cellular functions and subsequent cell death. Because PDI is non-selective and affects multiple cellular targets, development of resistance or tolerance to PDI has been considered to be unlikely and attempts to induce S. aureus resistance or tolerance upon repeated sub-lethal doses of PDI have not succeeded. However, multiple aspects of PDI suggest that development of tolerance is highly probable, in particular when PDI is used for treatment of infections where the environment at the infection site prevents penetration of PDI at a level sufficient to cause death of all bacteria and with tolerant phenotypes emerging from the surviving bacteria. In this study, we sought to identify the mechanisms that contribute to PDI tolerance in S. aureus. S. aureus HG003 and the isogenic HG003DmutSL strain with defects in DNA mismatch repair were used to evaluate the response of S. aureus and the roles of DNA mismatch repair and gene regulatory networks to repeated sublethal doses of PDI. Global transcriptome and genome analyses were used in an agnostic approach to identify the underlying transcriptional responses and genetic adaptations that occur as a result of repeated PDI and contribute to PDI tolerance. Our results reveal multiple metabolic, transport and cell wall biogenesis pathways that contribute to PDI tolerance, and a S. aureus regulatory gene likely responsible for the adaptive transcriptional response to PDI.

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Fluorescence Polarization of Methylene Blue as a Quantitative Marker of Breast Cancer at the Cellular Level.

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