Porphyrin-nanosensor conjugates : novel tools for the study of cellular response to oxidative stress

• Main Author: Josefsen, Leanne B.
• Other Authors: Boyle, Ross W.
• Published: University of Hull 2007
• Subjects: 571.9453; Chemistry
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.441585

The cellular environment is heterogeneous in nature and home to a multitude of species and reactions. If the labyrinth behind these reactions and signaling pathways (along with those leading to cell death) can be decoded it may be possible to identify mal-functioning and apoptotic signaling pathways. The particular interest for this Project is the correlation between reactive oxygen species (ROS) and altered calcium homeostasis. Oxidative stress has been linked to a number of disease states, including heart and Alzheimer's disease; although the exact mechanism(s) of induction are not fully understood. If the relationship between ROS and calcium perturbations can be elucidated further there is the potential to unravel disease "fingerprints". Conventional spectroscopic techniques have over the years provided insight into intracellular signaling events. However, limitations have been encountered with their toxicity and the difficulties associated with controlling their sub-cellular localisation and relative concentrations at these sites. Therefore, the aim is to create a more reliable tool which is capable of observing/monitoring a biologically relevant intracellular species; more specifically, tracking the intracellular response to oxidative stress via luminescent detection of the major signaling ion calcium. The strategy for achieving such a system/tool is based on the hybridisation of photosensitiser chemistry and polymeric nanoscience. A major advantage of this type of system/tool is the entrapment of a fluorescence species within an inert matrix of low toxicity and the in situ generation of ROS. The nanosensing component of the target system was successfully afforded by a free-radical emulsion polymerisation technique and conjugated (via a thiourea bond) to a water-soluble porphyrin. The ability of the target system to generate ROS and any transfer of energy between the two component chromophores was evaluated via time-correlated single photon counting (TCSPC). The potential of the system/tool to effect oxidative stress and chelate free intracellular calcium ions was assessed in human neuroblastoma and human embryonic cell-lines. Findings from the investigations suggested that the target porphyrin-nanosensor system is capable of generating singlet oxygen (in solution) and detecting free calcium ions (in solution and in vitro).