In this week’s WCMR ‘Research in Progress’ meeting we heard from Polly Usher, who told us about her PhD project that aims to develop a high throughput fluorescence screen to test potential therapies for mitochondrial disease. But what does this involve? Here, Polly tells us more.
Mitochondria are complex organelles whose healthy function relies on a lot of different factors. One of these is the mitochondrial redox environment. Redox is a term used to describe reactions which involve reduction and oxidation – or passing electrons between two molecules. The mitochondrial redox environment is made up of three pairs of molecules: NADP+ and NADPH, NAD+ and NADH, and FAD and FADH2. The idea behind redox is that each pair, such as NAD+ and NADH, exist together in a ratio. This means when NAD+ is turned into NADH, the opposite is happening somewhere else in the mitochondria. This makes sure the ratio is maintained. If you imagine a see-saw with NAD+ on one side and NADH on the other, the aim is to keep them balanced at all times.
Sometimes mitochondria lose this balance, either by a decrease in NAD+ levels (which can happen as we age) or through an increase in NADH (like when Complex I which forms part of the energy chain isn’t working properly). The main aim of my research project is to look for compounds which can rebalance the mitochondrial NAD+/NADH ratio, or improve Complex I activity, to try and treat mitochondrial disorders. To do this, I have been working with fluorescent cells and specialised analysis to detect levels of NAD+ and NADH within mitochondria. This means I can test a range of different compounds to see if any have the effect we’re looking for.
The video above shows how fluorescence can be used to show NADH levels changing within cells when you add a compound that blocks Complex IV of the energy chain. This results in a blockage along the energy chain which causes a back log of electrons, like cars in a traffic jam. When the blockage reaches Complex I it stops the energy chain from working. As you can see in the video, this leads to a sudden increase in NADH (brighter cells). As the blockage clears the fluorescence goes back to normal.