Hyperspectral imaging using intracellular spies : quantitative real-time measurement of intracelluar parameters in vivo during host-pathogen interaction

In the framework of pathogen-host cell interaction studies, programmed cell death has been appeared to be one of those cell events that are governed by different mechanisms. The ubiquitous fungus Aspergillus fumigatus is a widespread microorganism that based on the host’s immune deficiency can cause variety of diseases. The organism is capable of manipulating the host’s immune system by either induction or inhibition of apoptosis. Hyperspectral imaging is a technique based on combining the classical spectroscopy and conventional digital image processing. The technique is also well-suited for the quantitative real time analysis of mixed overlapping fluorescence signals transmitted from several sources within one sample. Therefore, a modified commercial fluorescence microscope; equipped with hyperspectral imaging technology, was gradually designed and developed to perform the kinetic studies of several intracellular infection-related events i.e. apoptosis or the measurement of pH in phagolysosome and cytosol at the single cell level. A significant relation between presence of melanin and the modulation of apoptosis as well as cell acidification was observed. By comparing different species of Aspergillus and its mutant strains, it is shown that this property is not only limited to A.fumigatus, but it also exists in the species that contain DHN-melanin i.e. Aspergillus clavatus and ∆abr2 mutant. The intracellular pH post-infection and during apoptosis was quantified and the pattern of intracellular pH changes upon infection was determined. The outcome would be versatile opportunities to benefit novel pharmaceuticals cell therapies i.e. in non-invasive therapeutic approaches and scheming the fate of apoptosis in cancer-cell researches.