Small molecule phosphorescent probes for O<sub>2</sub>imaging in 3D tissue models

Dmitriev, Ruslan I.; Kondrashina, Alina; Koren, Klaus; Klimant, Ingo; Zhdanov, Alexander V.; Pakan, Janelle M.P.; McDermott, Kieran W.; Papkovsky, Dmitri B.

doi:10.1039/c3bm60272a

Cited by 101 publications

(124 citation statements)

References 70 publications

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“…O 2 -sensitive probes MitoXpress®-Xtra [43], MitoXpress®-Intra NanO2 [44], Pt-Glc [45] and pH-sensitive probe pH-Xtra [46] were from Luxcel Biosciences (Cork, Ireland). Cytosolic pH probe BCECF AM, Lipofectamine® 2000 and Opti-MEM I were from Invitrogen Life Technologies (Carlsbad, CA).…”

Section: Methodsmentioning

confidence: 99%

“…The tissue was washed with Krebs buffer, covered with 2 mL of Krebs buffer containing 2% BSA and Pt-Glc probe (20 μM) and incubated for 3 h at 37°C in a CO 2 incubator. After washing, fresh Krebs buffer (3 mL) supplemented with 2 mM L-glutamine and 0.5 μM nifedipine (to prevent muscle contraction) was added and samples were imaged on a confocal PLIM system described in [45]. The system was based on an upright fluorescent microscope AxioExaminer Z1 with 20 ×/1.0 W-Plan-Apochromat water immersion objective (Carl Zeiss), DCS-120 confocal scanner, TCSPC module (Becker & Hickl) and R10467U-40 photon counting detector (Hamamatsu Photonics K.K.).…”

Section: Measurement Of Intracellular Omentioning

confidence: 99%

“…Measurement of iO 2 was performed using the phosphorescence quenching method [50] on Victor 2 plate reader (NanO2 probe) and confocal microscope (NanO2 and Pt-Glc probes) [45].…”

Section: Measurement Of Intracellular Omentioning

confidence: 99%

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A novel effect of DMOG on cell metabolism: direct inhibition of mitochondrial function precedes HIF target gene expression

Zhdanov

Okkelman

Collins

et al. 2015

Biochimica et Biophysica Acta (BBA) - Bioenergetics

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Abnormal accumulation of oncometabolite fumarate and succinate is associated with inhibition of mitochondrial function and carcinogenesis. By competing with α-ketoglutarate, oncometabolites also activate hypoxia inducible factors (HIFs), which makes oncometabolite mimetics broadly utilised in hypoxia research. We found that dimethyloxalylglycine (DMOG), a synthetic analogue of α-ketoglutarate, commonly used to induce HIF signalling, inhibits O2 consumption in cancer cell lines HCT116 and PC12, well before activation of HIF pathways. A rapid suppression of cellular respiration was accompanied by a decrease in histone H4 lysine 16 acetylation and not abolished by double knockdown of HIF-1α and HIF-2α. In agreement with this, production of NADH and state 3 respiration in isolated mitochondria were down-regulated by the de-esterified DMOG derivative, N-oxalylglycine. Exploring the roles of DMOG as a putative inhibitor of glutamine/α-ketoglutarate metabolic axis, we found that the observed suppression of OxPhos and compensatory activation of glycolytic ATP flux make cancer cells vulnerable to combined treatment with DMOG and inhibitors of glycolysis. On the other hand, DMOG treatment impairs deep cell deoxygenation in 3D tissue culture models, demonstrating a potential to relieve functional stress imposed by deep hypoxia on tumour, ischemic or inflamed tissues. Indeed, using a murine model of colitis, we found that O2 availability in the inflamed colon tissue rapidly increased after application of DMOG, which could contribute to the known therapeutic effect of this compound. Overall, our results provide new insights into the relationship between mitochondrial function, O2 availability, metabolic reprogramming and associated diseases.

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Section: Methodsmentioning

confidence: 99%

Section: Measurement Of Intracellular Omentioning

confidence: 99%

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A novel effect of DMOG on cell metabolism: direct inhibition of mitochondrial function precedes HIF target gene expression

Zhdanov

Okkelman

Collins

et al. 2015

Biochimica et Biophysica Acta (BBA) - Bioenergetics

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“…Fig. 19 demonstrates the capability of confocal TCSPC-PLIM, showing oxygen maps of (A) a resting 2D culture of MEF cells and (B) a single 2D slice from a PC12 multi-cellular 3D (spheroid) aggregate, both stained with a PtPFPP type porphyrin conjugated to four glucose residues (τ ¼ 57 μs) [86]. The sensitivity of the porphyrin conjugate (Pt-Glc) to changes in oxygen concentration in vitro is demonstrated by a series of lifetime maps recorded after stimulation with a variety of chemical treatments: (a) FCCP (activates mitochondrial respiration), (b) KCl (membrane depolarizing agent) and (c) sulfite (chemically deoxygenates the sample), known to alter cell respiration and therefore intercellular O 2 concentrations.…”

Section: Confocal Tcspc-plimmentioning

confidence: 98%

Time-Resolved Emission Imaging Microscopy Using Phosphorescent Metal Complexes: Taking FLIM and PLIM to New Lengths

Baggaley

Weinstein

Williams

2014

Luminescent and Photoactive Transition Metal Complexes as Biomolecular Probes and Cellular Reagents

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Luminescent metal complexes are increasingly being investigated as emissive probes and sensors for cell imaging using what is traditionally termed fluorescence microscopy. The nature of the emission in the case of second-and third-row metal complexes is phosphorescence rather than fluorescence, as it emanates from triplet rather than singlet excited states, but the usual terminology overlooks the distinction between the quantum mechanical origins of the processes. In steady-state imaging, such metal complexes may be alternatives to widely used fluorescent organic molecules, used in exactly the same way but offering advantages such as ease of synthesis and colour tuning. However, there is a striking difference compared to fluorescent organic molecules, namely the much longer lifetime of phosphorescence compared to fluorescence. Phosphorescence lifetimes of metal complexes are typically around a microsecond compared to the nanosecond values found for fluorescence of organic molecules. In this contribution, we will discuss how these long lifetimes can be put to practical use. Applications such as time-gated imaging allow discrimination from background fluorescence in cells and tissues, while increased sensitivity to quenchers provides a means of designing more responsive probes, for example, for oxygen. We also describe how the technique of fluorescence lifetime imaging microscopy (FLIM) -which provides images based on lifetimes at different points in the image -can be extended from the usual nanosecond range to microseconds. Key developments in instrumentation as well as the properties of complexes suitable for the purpose are discussed, including the use of two-photon excitation methods. A number of different research groups have made pioneering contributions to the instrumental set-ups, but the E. Baggaley and J.A. Weinstein Department of Chemistry, University of Sheffield, Sheffield S3 7HF, UK e-mail: liz.baggaley@sheffield.ac.uk; julia.weinstein@sheffield.ac.uk J.A.G. Williams (*) Department of Chemistry, Durham University, Durham DH1 3LE, UK e-mail: j.a.g.williams@durham.ac.uk terminology and acronyms have not developed in a systematic way. We review the distinction between time-gating (to eliminate background emission) and true timeresolved imaging (whereby decay kinetics at each point in an image are monitored). For instance, terms such as PLIM (phosphorescence lifetime imaging microscopy) and TRLM (time-resolved luminescence microscopy) refer essentially to the same technique, whilst TREM (time-resolved emission imaging microscopy) embraces these long timescale methods as well as the more well-established technique of FLIM.Keywords Cell imaging Á Fluorescence Á Fluorescence lifetime imaging microscopy Á Imaging Á Iridium Á Metal complex Á Microscopy Á Oxygen sensing Á Phosphorescence Á Phosphorescence lifetime imaging microscopy Á Platinum Á Ruthenium Á Time-resolved emission imaging microscopy Á Time-resolved fluorescence Á Time-resolved luminescence microscopy Á Tissue imaging

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“…1,4,11,12 As a result, advanced sensing platforms were developed to recognize various biologically relevant analytes, including (but not limited to) detection of cyanide ions, 13 biothiols 14 (including thiourea 15 ), b-galactosidase, 16 dopamine receptor, 17 or inhibitor of tumor necrosis factor-a. 18 Moreover, the sensitivity of emission parameters (intensity and lifetime) to oxygen concentration makes possible mapping of oxygen concentration in in vitro and in vivo experiments, 19,20 including quantitative measurements with the use not only emission intensity but also phosphorescence lifetime imaging (PLIM) techniques, [21][22][23][24][25] which provides considerably higher precision in oxygen biodistribution mapping. This turns traditional luminescence imaging into the instrument, which allows for functional analysis of physiological status of the targeted objects.…”

Section: Introductionmentioning

confidence: 99%

Water-soluble cyclometalated platinum(ii) and iridium(iii) complexes: synthesis, tuning of the photophysical properties, and in vitro and in vivo phosphorescence lifetime imaging

Solomatina

Lukina

et al. 2018

RSC Adv.

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This paper presents synthesis and photophysical investigation of cyclometalated water-soluble Pt(II) and Ir(III) complexes containing auxiliary sulfonated diphosphine (bis(diphenylphosphino)benzene (dppb), P^P*) ligand. The complexes demonstrate considerable variations in excitation (extending up to 450 nm) and emission bands (with maxima ranging from ca. 450 to ca. 650 nm), as well as in the sensitivity of excited state lifetimes to molecular oxygen (from almost negligible to more than 4-fold increase in degassed solution). Moreover, all the complexes possess high two-photon absorption cross sections

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Small molecule phosphorescent probes for O₂imaging in 3D tissue models

Abstract: PtPFPP-carbohydrate conjugates are promising O2probes for 3D PLIM imaging of live spheroids and brain explants.

Cited by 101 publications

References 70 publications

A novel effect of DMOG on cell metabolism: direct inhibition of mitochondrial function precedes HIF target gene expression

A novel effect of DMOG on cell metabolism: direct inhibition of mitochondrial function precedes HIF target gene expression

Time-Resolved Emission Imaging Microscopy Using Phosphorescent Metal Complexes: Taking FLIM and PLIM to New Lengths

Water-soluble cyclometalated platinum(ii) and iridium(iii) complexes: synthesis, tuning of the photophysical properties, and in vitro and in vivo phosphorescence lifetime imaging

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Small molecule phosphorescent probes for O2imaging in 3D tissue models

Abstract: PtPFPP-carbohydrate conjugates are promising O2probes for 3D PLIM imaging of live spheroids and brain explants.

Cited by 101 publications

References 70 publications

A novel effect of DMOG on cell metabolism: direct inhibition of mitochondrial function precedes HIF target gene expression

A novel effect of DMOG on cell metabolism: direct inhibition of mitochondrial function precedes HIF target gene expression

Time-Resolved Emission Imaging Microscopy Using Phosphorescent Metal Complexes: Taking FLIM and PLIM to New Lengths

Water-soluble cyclometalated platinum(ii) and iridium(iii) complexes: synthesis, tuning of the photophysical properties, and in vitro and in vivo phosphorescence lifetime imaging

Contact Info

Product

Resources

About

Small molecule phosphorescent probes for O₂imaging in 3D tissue models