Modeling the dynamics of hypoxia inducible factor-1α (HIF-1α) within single cells and 3D cell culture systems.

HIF (hypoxia inducible factor) is an oxygen-regulated transcription factor that mediates the intracellular response to hypoxia in human cells. There is increasing evidence that cell signaling pathways encode temporal information, and thus cell fate may be determined by the dynamics of protein levels. We have developed a mathematical model to describe the transient dynamics of the HIF-1α protein measured in single cells subjected to hypoxic shock.

Intracellular O2 sensing probe based on cell-penetrating phosphorescent nanoparticles.

A new intracellular O(2) (icO(2)) sensing probe is presented, which comprises a nanoparticle (NP) formulation of a cationic polymer Eudragit RL-100 and a hydrophobic phosphorescent dye Pt(II)-tetrakis(pentafluorophenyl)porphyrin (PtPFPP). Using the time-resolved fluorescence (TR-F) plate reader set-up, cell loading was investigated in detail, particularly the effects of probe concentration, loading time, serum content in the medium, cell type, density, etc. The use of a fluorescent analogue of the probe in conjunction with confocal microscopy and flow cytometry analysis, revealed that cellular uptake of the NPs is driven by nonspecific energy-dependent endocytosis and that the probe localizes inside the cell close to the nucleus.

Microdialysis based device for continuous extravascular monitoring of blood glucose.

Glycemic control of intensive care patients can be beneficial for this patient group but the continuous determination of their glucose concentration is challenging. Current continuous glucose monitoring systems based on the measurement of interstitial fluid glucose concentration struggle with sensitivity losses, resulting from biofouling or inflammation reactions. Their use as decision support systems for the therapeutic treatment is moreover hampered by physiological time delays as well as gradients in glucose concentration between plasma and interstitial fluid.

Evaluation of the derivates of phosphorescent Pt-coproporphyrin as intracellular oxygen-sensitive probes.

Several new derivatives of the phosphorescent Pt(II)-coproporphyrin (PtCP) were evaluated with respect to the sensing of intracellular oxygen by phosphorescence quenching. Despite the more favorable molecular charge compared to PtCP, self-loading into mammalian cells was rather inefficient for all the dyes, while cell loading by facilitated transport using transfection reagents produced promising results. The PtCP-NH(2) derivative, which gave best loading efficiency and S/N ratio, was investigated in detail including the optimisation of loading conditions, studies of sub-cellular localization, cytotoxicity, oxygen sensitivity and long-term signal stability.

Imaging of cellular oxygen and analysis of metabolic responses of mammalian cells.

Many parameters reflecting mitochondrial function and metabolic status of the cell, including the mitochondrial membrane potential, reactive oxygen species, ATP, NADH, ion gradients, and ion fluxes (Ca(2+), H(+)), are amenable for analysis by live cell imaging and are widely used in many labs. However, one key metabolite - cellular oxygen - is currently not analyzed routinely. Here we present several imaging techniques that use the phosphorescent oxygen-sensitive probes loaded intracellularly and which allow real-time monitoring of O(2) in live respiring cells and metabolic responses to cell stimulation.