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PubMed · 6189595

Fluorescent nitroheterocycles for identifying hypoxic cells.

P L Olive·R E Durand
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Abstract

Binding of several nitroheterocycles by mammalian cells is a function of the ambient oxygen concentration; anoxic single cells bind up to 10 times as much of these drugs as do aerobic cells. We thus hypothesized that fluorescent nitroheterocyles could be used to quantitate the fraction of hypoxic cells in multicell systems, and this was tested in multicell spheroids using a relatively nontoxic nitrofuran, trans-5-amino-3-[(5-nitro-2-furyl)vinyl]-1,2,4-oxadiazole. Binding of trans-5-amino-3-[(5-nitro-2-furyl)vinyl]-1,2,4-oxadiazole, as quantified by flow cytometry, was highly responsive to external oxygen concentration. To assess the relevance of the observed fluorescence, fluorescence-activated cell sorting was used to examine the radiosensitivity of cells as a function of their fluorescence intensity. The most fluorescent were indeed the most radioresistant (i.e., contained the least oxygen). Additional results confirm the general feasibility of using fluorescent nitroheterocycles as hypoxic cell probes but also reveal that cellular binding of these agents is not exclusively dependent upon cellular oxygen content.

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5-Amino-3-((5-nitro-2-furyl)vinyl)-1,2,4-oxadiazoleAnimalsCell LineCell SurvivalCricetinaeCricetulusFlow CytometryFluorescenceNitrofuransOxygenOxygen ConsumptionJournal ArticleResearch Support, U.S. Gov't, P.H.S.

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P L Olive, R E Durand. 1983. Fluorescent nitroheterocycles for identifying hypoxic cells.. https://pubmed.ncbi.nlm.nih.gov/6189595/

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5-Amino-3-((5-nitro-2-furyl)vinyl)-1,2,4-oxadiazol↗
PubMed1985

Fluorescent probes for cellular hypoxia: lack of transfer of fluorescence between cells in vitro.

Fluorescent nitroheterocycles may be useful as probes for cellular hypoxia. Reductive metabolism of AF-2 (cis 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide) and NFVO (trans-5-amino-3((5-nitro-2-furyl)vinyl)1,2,4-oxadiazole) results in intracellular accumulation of fluorescent molecules; the mean cellular fluorescence has previously been shown to be related to the cellular oxygen content during incubation in vitro. However, factors in addition to oxygen and nitroreductive activity may also affect cellular accumulation of these drugs. The stability of cellular fluorescence and possible diffusion of metabolites were examined by flow cytometric analysis of mouse and hamster fibroblasts exposed to NFVO and AF-2. Incubation of cells with 14C-AF-2 allowed calibration of the flow cytometer for AF-2 fluorescence; 5 X 10(8) molecules/cell resulted in double the spontaneous cellular fluorescence. Cellular fluorescence was stable for days after exposure to AF-2, and no evidence of transfer between exposed and unexposed cells was observed. For concentrations resulting in less than 5 X 10(9) AF-2 adducts/cell, all of the metabolites could be accounted for intracellularly. Therefore, it is unlikely that transfer of reduced nitroheterocycles occurs between cells.

5-Amino-3-((5-nitro-2-furyl)vinyl)-1,2,4-oxadiazol↗