Microscopy innovation offers cataract formation insights
Cataracts – a clouding of the lens of the eye – is a common age-related condition that can lead to loss of vision. Other contributing factors include genetics, solar UV light and smoking. “The eye lens is a very complex tissue,” notes Cata-rotors project researcher Petr Sherin from Imperial College London in the United Kingdom. “Unlike other tissues, the lens’ major components – proteins and lipids – do not renew over a lifetime. As a result, defects can accumulate, eventually altering the lens’ function and possibly leading to cataract formation.” While contributing factors have been identified, scientists do not know exactly what processes actually cause cataract formation, or whether there is any way to prevent or reverse these processes. “One theory behind the formation of cataracts is the appearance of a ‘diffusion barrier’,” explains Sherin. “This prevents the movement of nutrients to different parts of the lens in middle age.” Scientists however have been unable to confirm this hypothesis. The Cata-rotors project, which was undertaken with the support of the Marie Skłodowska-Curie Actions programme, sought to address this.
Identifying structural changes
To achieve its aims, Marie Skłodowska-Curie fellow Petr Sherin and the project coordinator Marina Kuimova applied a new microscopy method – developed in the Kuimova lab – called Fluorescence Lifetime Imaging Microscopy (FLIM). This was used in combination with fluorescent probes sensitive to viscosity, called molecular rotors. These rotors ‘light up’ the regions of cells that are more viscous (or gloopier) compared to other regions. This technique was applied to real eye lens samples, after natural ageing or after exposure to light or temperature (‘accelerated ageing’). The aim was to identify any differences in viscosity, which could offer support for the diffusion barrier hypothesis. “First, lens samples were sliced and stained with fluorescence rotors, making sure that the tissue was not damaged or dehydrated in the process,” says Kuimova. “This work was challenging and required high-level skills in tissue sectioning.” Next, large sets of FLIM images were recorded and analysed. “These provided us with detailed information on how viscosity was distributed within lens cells,” explains Sherin.
Looking for proof
Two key results were achieved. Visual changes in cells from central and peripheral regions of a lens in middle age were identified, and the differences in their viscosity recorded. “This is one of the most direct proofs of ‘diffusion barrier’ formation available to date,” adds Sherin. “To directly confirm this finding however, we need to accumulate more statistically significant data.” The project also found that the membranes of porcine eye lens cells become considerably less stiff (more fluid) upon exposure to solar UV light. This unexpected result may act as a compensatory mechanism for increased lens stiffness in elderly lenses, by providing a minimally required flow of nutrients. “Above all, the project demonstrated the applicability of this ‘FLIM – molecular rotor’ approach to visualising important properties within eye lens tissues,” says Kuimova. “Such information could be used as the basis for developing new therapeutic cataract treatments. Surgery is currently the only option.” The next major step forward is to better understand the molecular mechanism behind the stiffness changes. This will involve identifying which chemicals and which reactions are involved, and hopefully offer options for medical intervention other than surgery.
Keywords
Cata-rotors, eye, cataract, microscopy, FLIM, lens, ageing, molecular
