Periodic Reporting for period 2 - LOGIC LAB (Molecular logic lab-on-a-vesicle for intracellular diagnostics)
Okres sprawozdawczy: 2020-11-01 do 2023-10-31
Such tailor-made diagnostic tools are urgently needed both in fundamental research and clinical routine. As a proof-of-concept, LogicLab will develop a molecular logic sensor platform to diagnose endothelial dysfunction (ED). A dysfunction of the the cells lining our blood vessels is the primary cause of many lifestyle related diseases such as atherosclerosis, which account according to the WHO for 60% of all deaths worldwide in 2005. In atherosclerosis – a chronic medical condition that can remain undetected for decades - so called plaques deposit at the walls of blood vessels leading to stenosis and eventually to serious clinical events such as heart attack or stroke. An early-stage diagnosis of atherosclerosis would allow for intervention long before a high and thus dangerous degree of stenosis is reached. By detection of key biomarkers for ED, such as low wall shear stress and low nitric oxide (NO) concentration, LogicLab aims to develop an analytical platform to diagnose ED and the diseases it possibly provokes at a primary level. The objective of LogicLab is to explore a new concept of molecular logic sensing that can be applied in a biological environment. Not till then, molecular logic sensors will find wide-spread and important application such as in intracellular sensing, critical care diagnostics and high-throughput screening. The approach of LogicLab is a stepwise transfer of the concept from solution to membrane models to real biological environments such as cells and tissue.
All newly synthesized compounds have been spectroscopically and theoretically characterized for their optical and electronic properties in solution in order to tune them for application in a biological environment. In parallel, stable vesicles and liposomes have been prepared and evaluated regarding their stability in buffer. These vesicles have been functionalized with photosensitzers and annihilator molecules and TTA-UC within the lipid membrane of the vesicle has been demonstrated. This serves as a proof of concept on the way to supramolecular logic sensing. Moreover, the functionalized vesicles have been demonstrated to fuse with two-dimensional (2D) models of the cell membrane. Cell cultures and co-cultures of different endothelial cell lines have been cultivated both in classical petri dish cell cultures and in microfluidic cell cultures.These 3D cell cultures cultivated inside a microfluidic channel represent a model of an artery and will be used to evaluate the supramolecular logic gates and molecules which release specific doses of nitric oxide upon light irradiation. To implement a therapeutic function into the vesicles, a light-activatable compound (photoNORM) for release of therapeutic amounts of nitric oxide has been synthesized, spectroscopically characterized and evaluated towards light-triggered NO release.
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