Australian Synchrotron X-ray and infrared imaging techniques have been used in a powerful combined approach to characterise the composition of amyloid plaques that are associated with Alzheimer’s disease. 

Alzheimer’s disease is major international health problem that accounts for 50-75 per cent of all cases of dementia in Australia. More than 400,000 Australians are living with dementia and it is the second leading cause of death.

Amyloid plaques are complex protein fragments which accumulate between nerve cells in the brain and may destroy connections between them, and are hallmarks of Alzheimer’s disease. 

“However, it is still not known if the plaques cause Alzheimer’s or whether the Alzheimer’s causes their formation, which is why we need to improve our understanding of protein structures within plaques, and the molecular and elemental composition of tissue surrounding the plaques“ said Dr Mark Hackett of Curtin University, who led the research. 

The study was published earlier in the year in Biochemistry.

As very few methods provide sufficient chemical information to study the composition and distribution of the plaques in excised tissue, the investigators decided to combine Synchrotron spectroscopic techniques with additional imaging methods, Raman spectroscopy and fluorescence microscopy.

“It is something that really hasn’t been done before in Australia and demonstrates the power of the approach” said Australian Synchrotron instrument scientist Dr David Paterson. He and Dr Mark Tobin of the Synchrotron were among a large team of collaborators from Curtin University, the University of Saskatchewan and The University of Adelaide.  

Histology, FTIR, XFM, and tissue autofluorescence imaging of Aβ-plaques

Metals have long been associated with amyloid plaques and Alzheimer’s, and a number of leading international research groups have used synchrotron techniques to reveal metal distribution within plaques. However, the exact role of metals in Alzheimer’s disease is still not known, “which is why it is important to correlate metal concentration and distribution within plaques to alterations of important biochemical parameters, such as lipids and proteins” said Paterson, who assisted with the collection and analysis of X-ray fluorescence microscopy data (XFM). 

The bright source of X-rays produced by the Australian synchrotron is a major advantage for XFM.

“You have a high energy X-ray coming in and if it is absorbed by an iron atom it will re-emit X-rays at a very specific energy and we have detectors that can tell the difference between the X-rays coming from iron or copper. The more iron atoms there are in a particular location, the more fluorescence we will see from there,” said Paterson.

Not only can XFM differentiate between different elements, but X-ray fluorescence microscopy is a direct imaging technique that does not involve any staining. This is really important, as the typical staining methods often used to study Alzheimer’s disease, may remove important chemical information from the tissue.

“To be able to study metal and molecule distribution, without staining, is a really unique capability, and is made possible with synchrotron light” said Hackett.

The XFM beamline was used to complement Raman spectroscopy and infrared microspectroscopy in determining the location of specific metals within the plaque and classes of molecules such as lipids, cholesterol and aggregated protein. The results indicated that intense zinc and some iron were found within the plaque core, while the copper is spread out in a cloud-like shape at the periphery.”

“Because you can overlay the fluorescence images of each element, you acquire a useful chemical composite of the plaque,” said Paterson.

Supporting the XFM data, infrared imaging and Raman microscopy provided crucial information about the molecular structure within the plaques, in this case, the presence and amount of lipids. Surprisingly, while aggregated proteins were found to localise with Zn and Fe in the plaque core, lipids were found to localise with Cu at the plaque periphery.

 “At this stage, we are unsure of the exact meaning of the co-localisation of lipids and Cu in the plaque core, however, we now have an imaging methodology that allows us to study this in the future, which is an important step forwards”, said Hackett.

“The IR microspectroscopy, when complemented by Raman microspectroscopy indicated that there was an increase in the levels of copper and  lipids at the periphery of the plaque,” said instrument scientist Dr Mark Tobin, who assisted in the data collection and analysis of the Synchrotron source FTIR.

 “Future research into the interactions between copper and the lipids in the amyloid plaque is worthy of further investigation.”  said Hackett.

http://pubs.acs.org/doi/pdf/10.1021/acs.biochem.7b00262

Approximately 190 participants attended the first combined ANSTO User Meeting, which was held at the Australian Synchrotron 22-24 November.

 

 

 

The event brought representatives of research communities together who have accessed various ANSTO infrastructure platforms.

 

 “It was an opportunity to look at the scientific challenges and questions that are being addressed and consider how multiple techniques and experimental methods can be applied to answering those questions,” said co-convenor Dr Miles Apperley, Head of Research Infrastructure, who spoke at the opening.

 

ANSTO has nine research infrastructure platforms in total, including the Australian Centre for Neutron Scattering and the Australian Synchrotron that provide user-focused open-access support to researchers from Australia and across the globe.

 

Plenary speakers included leading Australian and International researchers.  

 

Dr Michael Drakopoulos reviewed the basics of using the I12 beamline at the Diamond Light Source in the UK.

 

Professor Ian Gentle Deputy Executive Dean and Associate Dean Research of the Faculty of Science at the University of Queensland spoke about the use of neutrons to understand diffusion and stability of materials in organic optoelectronic devices.  

 

Professor Elena P. Ivanova, a nano/biotechnology expert from the University of Swinburne, shared her research deciphering the enigma of bactericidal pattern of insect wing epicuticle.


Associate Professor Rachel Popelka-Filcoff of Flinders University shared how Indigenous Australian natural pigments can be explored using nuclear and spectroscopic methods.

 

Professor Sharon Robinson, Associate Dean Graduate Research and a Senior Professor in Biological Sciences discussed species on the move in Antarctic Terrestrial Communities.

 

The speakers emphasised emerging science trends in their respective fields and how new technologies and capabilities benefit that research.

 

Both ANSTO and researchers from other Australian institutions contributed to sessions over the three days.

 

ANSTO scientists described the capabilities that support their own research outcomes as they develop and apply leading edge instrument methods and techniques.

 

They included include Tom Carodoc- Davies, Gabriel Murphy, Jason Price, Kathleen Wood, Katie Sizeland, Pimm Vongsvivut, Wai Tung Lee, Mark Tobin, Vladimir Levchenko,

Anna Sokolova, Cameron Kewish, Anton Stampfl, Daniele Hausermann, Susan Cumberland, Santosh Panjikar, Michael Hotchkis, Jun Aishima, Peter Kappen, Dehong Yu, Stephen Holt, Ulf Garbe, Garry McIntyre, Zeljko Pastuovic, Mark Reid, and Zhiyang Wang.

 

Prof Michael James outlined the new BRIGHT Beamlines that are planned for the Australian Synchrotron.

 

Prof Andrew Peele awarded the Stephen Wilkins Medal to Leonie van ‘t Hag for her PhD thesis on the application of a crystallographic technique that is used to crate 3d structures of proteins.

 

“It is not just stainless steel and digital systems that make up the infrastructure which our users encounter at our facilities, but the expertise and dedication of our staff who ensure that they can turn that information into knowledge and innovation,“ said Apperley.

  

The organising committee was chaired by Dr Anthony Chesman from CSIRO, who is a user of the Australian Synchrotron, and comprised representatives of the ANBUG and Synchrotron user communities and ANSTO platforms.

 

The combined Users Meeting will be held in the multi-disciplinary format every second year and revert to separate user community meetings on the alternate years to ensure important focused and discipline specific work can continue.

PHD student Dr Leonie van ‘t Hag has been awarded the prestigious 2017 ANSTO, Australian Synchrotron Stephen Wilkins Medal for her PhD thesis.

 

The award recognises her research to improve the method to crystallise proteins and peptides in order to study their structure, using a technique called crystallography.

 

“Leonie’s insights into crystallisation processes could significantly help the development of treatments for a variety of illnesses,” said Australian Synchrotron Director, Professor Andrew Peele.

 

Most solid material in the world is made of crystalline structures. Crystals are made up of rows and rows of atoms or molecules stacked up like boxes in a warehouse, in different arrangements.

 

The science of determining these atomic or molecular structures from crystalline materials is called crystallography.    

 

 

 

 

The Stephen Wilkins Medal is awarded annually to a PhD student at an Australian or New Zealand University who is judged to have completed the most outstanding thesis of the past two years.

 

Professor Peele said Dr van ’t Hag, who while working at the University of Melbourne, CSIRO and RMIT University was a prolific user of ANSTO facilities at the Australian Centre for Neurtron Scattering, the National Deuteration Facility and the Australian Synchrotron, was thoroughly deserving of her medal.

 

The type of crystallisation explored by Dr van ‘t Hag  is one of a number of methods used to create 3D structures of proteins, including membrane proteins that are typically very difficult to crystallise.

 

“The application of this technique can lead more success in solving the 3D structures of proteins, which are important for rational drug design for a wide range of diseases,” Professor Peele said.

 

“I congratulate Dr van ’t Hag for her work in this field and on behalf of the Wilkins family and ANSTO. A contribution of $3000 will also be awarded to support her further career development.”

 

Dr Mike James, who is the Head of Science at The Australian Synchrotron, congratulated Dr Leonie, and said she was most deserving of the award and the recognition.

 

“Steve Wilkins was a friend and mentor to many young researchers, so it’s fitting the ANSTO, Australian Synchrotron annual thesis medal be named in his honour,” said Dr James.

 

“Leonie is certainly a very worthy recipient, and we are very excited to see where she will go with her career, and what benefits and impacts the application of her research will bring.”

 

The award is named in honour of Stephen (“Steve”) Wilkins, a widely-respected and internationally-renowned X-ray scientist, who made pioneering contributions in many areas of X-ray science and optics. 

 

To be eligible, work is required to be undertaken at and acknowledge, the Australian Synchrotron, or work undertaken at international synchrotron facilities and funded by various ANSTO access programs.

 

Dr Leonie van ‘t Hag will be attending the event presentation ceremony to present her thesis and receive the award from Mrs Linda Wilkins – wife of the late Stephen Wilkins.
 

Media contact: Phil McCall 0438 619 987