The not so powerful omega-3

Omega-3 fatty acids have been in the media a great deal over recent years. They can lower our risk of heart disease and they may even have neuroprotective properties (for example limit damage to the brain and spinal cord after acute injuries).

But, what about in MND? Could this dietary supplement have an effect?

MND Association-funded researchers have found out, rather unexpectedly, that the omega-3 fatty acid eicosapentaenoic acid (EPA) actually accelerates disease progression in an animal model of MND which is based on a SOD1 mutation, when EPA is given before symptoms first appear (this is sometimes known as the pre-symptomatic stage).

Why omega-3 might have had an effect

Omega-3 polyunsaturated fatty acids are natural compounds primarily found in oily fish such as sardines, mackerel or salmon. They have been widely associated with significant health benefits and researchers have reported that some long-chain polyunsaturated fatty acids may be beneficial in several neurological conditions.

Previous research in rats has shown that dietary food supplements, containing omega-3 long-chain polyunsaturated fatty acids (including EPA), reversed age-related problems in neurones (nerve cells) and also enabled the growth of new neurons.

The neuroprotective properties of EPA could occur through a variety of mechanisms such as reducing oxidative stress (damage due to low oxygen levels), reducing neuroinflammation  and the activation of anti-‘cell death’ pathways. These are all factors that are relevant in MND.

A number of studies have found that high blood lipids (the breakdown product of dietary fats) are a common feature in ALS (the commonest form of MND), and are correlated with increased survival. High-fat diets have been studied in the lab to further investigate this and have been shown in mice to delay motor neurone death and extend lifespan.

What the researchers found

Due to these previous studies the researchers decided to study one omega-3 long-chain polyunsaturated fatty acid in particular, EPA, to assess whether it had neuroprotective effects in a mouse model of ALS based on a mutation in the enzyme SOD1.

Mouse models are commonly used to study the causes of the disease and investigate potential treatments in MND. A SOD1 mouse model is a mouse that has been given a faulty MND-causing gene producing an enzyme known as ‘SOD1’, which is known to cause 20% of cases of the rare inherited form of MND.

The researchers intended to study the effects of dietary EPA when given at disease onset (the symptomatic stage when symptoms first appear) or at the pre-symptomatic stage.

The mice were fed either a standard rodent powdered diet (control) or a diet supplemented with EPA-enriched oil. The researchers then looked at a number of factors such as: disease progression, survival and body weight to find out if there were any differences.

When a diet supplemented with EPA was given at the symptomatic stage there was no significant difference in the development of MND compared to the control group (mice who were fed a standard rodent powdered diet). However, rather unexpectedly, when the EPA diet was given at the pre-symptomatic stage, the researchers found that the diet accelerated the progression of MND, but did not affect disease onset.

Glial cells (such as astrocytes and microglia) were also affected, and found in reduced levels when the mice were given the EPA diet.

Overall, the researchers found that long-chain omega-3 fatty acid EPA-enriched diets have no impact on disease onset or survival. Unexpectedly, if dietary EPA is given before symptoms appear it can actually accelerate the progression of MND.

What this means for people living with MND

The omega-3 fatty acid EPA, although it may have other health benefits, appears to have the potential to be more damaging rather than protective in this specific MND mouse model. The results from this study have highlighted the need for caution by those who are at risk of developing MND, who may use these long-chain omega-3 fatty acids dietary supplements, which are freely available, over prolonged periods of time.

This study has shown that individuals who carry the SOD1 inherited form of MND in particular should take extreme caution with diets enriched in long-chain omega-3 fatty acids such as EPA. For the future, it remains to be seen if EPA has also negative effects in other models of MND (eg zebrafish or flies with the C9orf72 mutation).

Dr Adina Michael-Titus (Blizzard Institute, Queen Mary University of London), one of the researchers involved in the study, commented “The most important point, in my view, is to be aware that we do not have yet the scientific evidence to prove that EPA or any other omega-3 fatty acids are dangerous for all forms of MND. The only experimental evidence we have so far is for a particular SOD1 mutation which leads to this disease (where the faulty SOD1 mutation is greatly overexpressed). More work is required and future research will help us fully assess and understand the potential or the risk associated with omega-3 fatty acids in people living with MND.”

Dr Andrea Malaspina

Dr Andrea Malaspina (a member of our Biomedical Research Advisory Panel) also commented on the results. “EPA was found to accelerate the progression of MND when given at the pre-symptomatic stage in a SOD1 mouse model. To fully assess the potential risk of EPA further research is needed in other animal models, with different MND mutations (as different mutations cause different metabolic changes) to see if a similar effect is observed. At present we can only say that EPA accelerates the progression of MND in a SOD1 mouse model and it is not known whether it accelerates the progression of all forms of MND.”

For more information about the rare inherited form of MND please see our website

References: Yip PK, Pizzasegola C, Gladman S, Biggio ML, Marino M, et al.  (2013) The Omega-3 Fatty Acid Eicosapentaenoic Acid Accelerates Disease Progression in a Model of Amyotrophic Lateral Sclerosis. PLoS ONE 8(4): e61626. doi:10.1371/journal.pone.0061626

The C9orf72 mystery begins to unravel even more of its secrets

In 2011 an international team of scientists, including three MND Association-funded researchers, identified the elusive C9orf72 gene located on Chromosome 9. Since this ground-breaking discovery, researchers from around the world have been trying to find a way to open-up and reveal more about this MND-causing gene.

Determined to get inside and unravel the secrets behind C9orf72, the Association is funding a number of new and exciting research projects to help solve the mystery. These projects look at, not one, but a number of different aspects to try and understand more about C9orf72.

In order to solve this mystery our C9orf72 researchers are following the clues using zebrafish, mice, flies and DNA samples.

How the C9orf72 MND mystery began

We each contain copies of 23 pairs of chromosomes, including the X and Y sex chromosomes. These chromosomes contain thousands of genes that portray our characteristics such as hair and eye colour. These genes are made up of DNA which can either be ‘coding’ to make a protein, or ‘non-coding’. For details of how genes make a protein see our earlier blog post.

Before C9orf72 was identified researchers had focused on an area on Chromosome 9 that appeared to be connected with both the rare inherited form of MND and the related neurodegenerative disease frontotemporal dementia (FTD).

Using a number of cutting-edge techniques the international team isolated the C9orf72 gene expanded GGGGCC hexanucleotide repeat as being a crucial player in both inherited MND and FTD. Not only did the researchers find a link between MND and FTD, they also found that C9orf72 was found in approximately 40% of cases of inherited MND (where there is a strong family history). This means that we now know 70% of the genes that cause the rare inherited form of MND. For more details on C9orf72 see our earlier blog post.

For more information on inherited MND please see our website.

So, researchers found C9orf72. The next question was ‘What does it do? Is the gene defect repeat itself, or the protein it makes responsible for causing MND? And what goes wrong in MND?’

Following the clues to solve C9orf72

Two recent research clues

Since 2011 researchers have been trying to answer these questions and find out more about C9orf72. This has led to a dramatic increase in research, including two papers published in February and March this year!

Prof Christian Haass (Munich Centre for Neurosciences, Germany), who recently presented at our 23rd International Symposium on ALS/MND in December 2012, published a paper on the 7 February in the journal Science. The second paper lead by Prof Leonard Petrucelli (Mayo Clinic, USA) was published open access in the journal Neuron on the 20 February.

In a big surprise, both researchers found that the presumed ‘non-coding’ C9orf72 GGGGCC repeat expansion actually made a protein. Normally these ‘non-coding’ regions do not make proteins so this was a very big surprise indeed!

The researchers found that these proteins formed large clumps in the brains, and throughout the central nervous system (CNS), of people with C9orf72 MND and/or FTD. Importantly, they did not find these clumps in healthy individuals or those with other neurological disorders.

It is currently unknown as to whether these protein clumps are involved in MND and/or FTD, but they may be a potential biomarker or a therapeutic target in this most common type of MND. The next step is for the researchers to find out whether these proteins actually cause MND and/or FTD.

Finding more evidence to piece together the clues

In addition to these two papers looking into the mystery behind C9orf72, the Association is funding some exciting new research projects, each looking at different things, to further understand more about this gene.

Dr Johnathan Cooper-Knock, MRC/MND Association Lady Edith Wolfson Clinical Research Fellow

Dr Johnathan Cooper-Knock (Sheffield Institute for Translational Neuroscience, UK) is already trying to identify how C9orf72 causes MND by utilising a genetic technique known as gene expression profiling. He is using samples from the Association’s DNA bank which are positive for the C9orf72 genetic mistake. Gene expression profiling is a technique which allows researchers to understand how the activity of genes contributes towards causing MND. (Traditional genetic studies are designed to look at which genes are affected, rather than their activity – ie when and how). Read more about Johnathan’s project here.

Developing new disease models enables us to understand the causes of MND and to test new therapies. One way to understand the function of C9orf72 and how this goes wrong in MND is to create a model. Our current research projects are developing new C9orf72 models in flies, mice and zebrafish.

Dr Frank Hirth (Kings College London, UK) will be producing a fly model, Dr Javier Alegre Abarrategui (University of Oxford) will be making a mouse model and Dr Andrew Grierson (University of Sheffield, UK) will be creating a zebrafish model.

All of these models aim to understand the function of C9orf72 and what goes wrong. The researchers hope to study what happens in MND and how this occurs by looking at behaviour and what happens when C9orf72 is ‘switched’ on and off. For more information about these exciting research projects please see our website.

Solving the mystery

All of our C9orf72 Association-funded research projects are using different approaches to look at C9orf72 in different ways as we are still unsure whether the protein or the repeat is the problem. From mice to flies all of these research projects together are helping to solve the mystery of C9orf72 and MND.

With the proteins formed by C9orf72 likely to be a potential biomarker or therapeutic target the two recent papers are adding to the growing number of clues, pointing researchers in the right direction to unravelling and solving the secrets of C9orf72.

References:

Mori, K. et al. The C9orf72 GGGGCC Repeat Is Translated into Aggregating Dipeptide-Repeat Proteins in FTLD/ALS. Science. 339(6125): 1335-1338. 2013 DOI: 10.1126/science.1232927

Ash, P. E. A. et al. Unconventional Translation of C9ORF72 GGGGCC Expansion Generates Insoluble Polypeptides Specific to c9FTD/ALS. Neuron. 77(4): 639-646. 2013 DOI: 10.1016/j.neuron.2013.02.004

Baking with proteins, mRNA and DNA

Each and every one of us is made up of thousands of different ingredients, which all combine together to create something amazing; life. Perhaps the most important of these are proteins.

Each protein in the body has its own special job to do. From making our muscles contract to controlling blood sugar, proteins are an essential ingredient in life.

In MND research we have identified a number of MND causing genes. These are genes that are found to be mutated in some people living with MND, which somehow causes the motor neurones to die. But, how does this happen? How does a gene form a protein? This blog post explains how an MND causing gene becomes a protein.

As simple as baking a cake

Here at the MND Association we love our cake. So, I thought what better way is there to describe how we make proteins?

cheesecake

Every cell in our body contains 23 pairs of chromosomes (46 in total), except for the egg and sperm cells that contain 23 chromosomes each.

Like a recipe book, these chromosomes hold all of our genetic material in the form of genes, in which everyone inherits two sets of (one from each parent).

Humans have approximately 24,000 genes, which each consist of their own DNA recipe to make a protein. Like cakes, proteins come in a range of different shapes and sizes, that come together to create you and me.

These DNA recipes are read by different cells to create the right protein for the job. For example, you would only make a wedding cake for a wedding, and the type of cake (chocolate or fruit) would depend on the wedding couple.

This is what happens in nerve cells (or motor neurones). A motor neurone will make a specific type of protein to help it grow, or to help it survive in low oxygen levels.

Following the recipe

A nerve cell creates a protein by finding the exact DNA recipe amongst the genes within the cell’s control centre, known as the nucleus. Once the recipe has been found the cell has a problem… The nucleus does not have the right tools to make a protein! The cell instead needs a specialised machine, or food mixer, which is only found outside of the nucleus called a ribosome.

In order to make the protein the DNA recipe needs to travel from the nucleus to the ribosome and this is done by means of a messenger. The DNA recipe can’t leave the nucleus so the cell ‘copies’ it into a messenger version, called mRNA.

The cell does this by removing certain parts of the DNA that do not affect the finished protein which are known as introns or ‘non-coding DNA’. This is known as ‘RNA splicing’ and is the same as removing raisons from a fruit cake. The cake is still made and still contains fruit, but the raisons are not essential in the finished cake.

mRNA can then travel the DNA recipe safely from the nucleus to the ribosome, where it can be finally made into a protein. Once made, this protein can then go on to do its specific job role (or in cake terms, be a wedding cake!).

ruined cake

Changing and ruining the recipe

Sometimes the DNA recipe in our genes can change through means of a mutation. Most of these are harmless spelling mistakes (sugarr instead of sugar) that do not affect the finished protein. However, sometimes these mutations can be so big and harmful (salt instead of sugar) that they do.

These kind of mutations are so big that the size, shape and structure of the protein can be changed – meaning that the protein can no longer do the job it was designed to do (our wedding cake is now no longer sweet and tasty, but ruined and salty!)

This is what happens in some of the MND causing genes. A big mutation occurs in the DNA recipe in a specific gene that causes the structure and shape of that protein to change. This change can then cause the proteins to ‘clump’ together in the motor neurones as they can no longer do the job they were designed to do.

Bake MND history

Expert bakers

An understanding of genes and how proteins are connected is essential for understanding how they can go wrong in MND. The Association funds a number of exciting research projects investigating the MND causing genes, along with the proteins they form.

To help raise awareness of MND you can bake your own cake as part of our ‘Bake it!’ fundraising campaign. For more information and to request a fundraising pack please see our website.

Major new research finding raises some old questions

Hot on the heels of ‘Brain Awareness Week’, comes ‘National Science Week’, with the University of Sheffield enthusiastically organising a numerous activities in their week-long Festival of Science and Engineering, including today’s Open Day at the Sheffield Institute of Translational Neuroscience (SITraN). This event was to include talks by Dr Chris McDermott and Prof Pam Shaw on MND and the role that SITraN plays in the search for effective treatments for neurodegenerative disease.

Unfortunately…… last night’s snow has put the kibosh on that, so instead of heading up the M1 to Sheffield, I decided to use some of the saved time to catch up on some reading – in particular a recent paper that came out in the journal Nature, from an international consortium, led by the scientists from the Austrian Academy of Sciences in Vienna.

The best seven pages in ten years

A senior MND researcher emailed me to say it’s one of the best papers he’s read in the past 10 years and I can understand where he’s coming from. Not only does the research identify a previously unknown cellular process that causes selective motor neuron degeneration, but it also appears to tie together several of the pieces of the pathological jigsaw: disruption of RNA metabolism, oxidative stress and programmed cell death pathways.

As impressive is the sheer amount of work that has gone into this seven-page paper. OK, there are also several extra pages of online supplementary material (one of the great benefits of online publication) but I reckon there is the equivalent of at least three PhD theses and several years of work in there!  

In a nutshell, the researchers created a mouse that has a defect in an enzyme called CLP1 and these mice develop progressive motor neuron degeneration. I’m not going to go into the detail, but rather focus on one interesting item that was buried in the text.

Genetic environment matters for CLP1

When researchers initially tried to create the mice they found that the mice all died well before birth. So they tried using a different strain of mice, but got the same result.

A third strain produced live mice, with normal numbers of motor neurons at birth. However from about the age of four months, these mice then developed a progressive muscle weakness and loss of motor neurons over the course of several months.

The paper focuses in on what’s going on in these mice, but it also raised additional questions for me, such as:

“Why did these mice survive into adulthood, when two other mouse strains didn’t – and is there something different in the genetic make-up of these mice that has basically protected them into adulthood rather than killing them as embryos?”

MND Association funded research on genetic environment

Other groups have noticed that when SOD1 mice are bred on different background strains, it can have a profound effect on disease progression and survival. This brings us nicely back to SITraN, as Prof Shaw and her colleagues are looking at precisely this issue, in an MND Association-funded collaboration with Prof Caterina Bendotti in Milan.

They are looking at the gene expression profiles (basically which genes are switched on and off) in the motor neurons of two strains of SOD1 mice, one of which develops the disease and later age and also lives much longer.  By working out patterns that are linked to specific biological processes, they are starting to pinpoint pathways which are driving the disease and also which ones might be slowing the disease. Some of their findings were presented at the most recent International Symposium (Abstract C61).

If there are protective genes at work in the mice, might the same be happening in humans?

The search for ‘good’ genes hots up

I’m often asked about Steven Hawking – how come he’s lived so long?  For years, one of my pet theories has been that there is something in his genetic make-up that didn’t stop the disease from occurring, but is ‘pushing back’. That’s becoming an increasingly popular and productive area of investigation – as genetic researchers extend their focus from finding ‘bad’ genes that cause or predispose people to develop MND, to potentially ‘good’ genes that might slow down the disease. A couple of candidates have been identified, most notably the EphA4 gene.

The search for these disease-modifying genes needs joined up collaboration between researchers around to world and it’s heartening to see how everyone in the field is starting to get together to pool their samples and data, which will allow the genetic profiles of those with exceptionally slowly progressing MND to be analysed in much larger numbers than ever before. If good genes can be identified and their roles understood, it will open up exciting new treatment opportunities.

Zebrafish show that ‘connector neurons’ are the key in early stages of MND

A recent study by Motor Neurone Disease Association-funded researcher Dr Tennore Ramesh from the Sheffield Institute for Translational Neuroscience (SITraN) has shown that even before the symptoms of MND occur, at the earliest stages of the disease, ‘connector neurones’ known as interneurons are already becoming damaged in the zebrafish.

Dr Tennore Ramesh

Zebrafish are ideal models for helping scientists understand what happens in MND. Unlike mice and fly models, zebrafish have transparent embryos which enable scientists to get a unique view of the developing neurones under a microscope! Scientists can also look at disease progression in adult zebrafish by looking at muscle strength and measuring their progress swimming against a current.

Not only are zebrafish useful for helping scientists understand what happens in MND, they are also an ideal drug screening model. Zebrafish and humans are more similar than you may think (see Kelly’s post) and potential new MND drugs can be screened quickly. Looking at how MND progresses in the zebrafish, before symptoms appear, can help us gain a better understanding of what causes the disease.

Motorways, dual carriage ways and slip roads

No, I’m not writing about travel alerts or the latest roads disruptions due to flooding or snow. In fact, these road systems happen to be a perfect example of what interneurons are, how they relate to motor neurones and what goes wrong in MND.

Our body consists of two types of motor neurones, which are known as upper and lower motor neurones. The upper motor neurones are found in the motor region of our brain and connect to the spinal cord. The lower motor neurones are found between the upper motor neurones in the spinal cord and connect to the muscles (e.g. in the arms and legs). Interneurons are the vital connections between the upper and lower motor neurones.

Interneurons are the ‘slip roads’ between upper and lower motor neurons

When a signal is sent from our brain to bend an arm it starts by travelling down an upper motor neurone. The signal then travels to a lower motor neurone via an interneuron. When the signal from the lower motor neurone reaches the muscle in our arm it causes the muscle to contract and bend.

In MND these upper and lower motor neurones become damaged and they are unable to transport the nerve signal from the brain to the muscle in our arm. This means we are unable to contract and bend, even though the brain is telling it to.

­­­In our road system scenario the upper motor neurones are the motorways (e.g. the M1), and the lower motor neurones are the dual carriageways that link the motorways to nearby towns (e.g. the A38). In order for an upper motor neurone to send a signal (e.g. a car) to a lower motor neurone it needs to go via an interneuron, which in our road system scenario is a ‘slip road’ – making these interneurons vital connections between motor neurones.

This study has given us a better understanding of what happens in MND at the early stages of the disease (before symptoms occur). The researchers found that interneurons became damaged before the motor neurones themselves. Therefore this shows that interneurons are important in the early stages of the disease and scientists can begin to look at ways of preventing interneuron damage to see whether this has an effect on MND.

Adding more evidence to the puzzle

This study showed that, in zebrafish, interneurons are involved in the early stages of MND, which adds further evidence to previous work by another MND Association-funded researcher. Dr. Martin Turner (Oxford) also found damaged interneurons at the early stages of the disease before symptoms of MND occur in humans, with other studies showing interneuron damage in SOD1 mice models.

The next step would be to look at ways of preventing these interneurons from becoming damaged, to see whether this has any effect on the progression of MND.

This research is the first article we have paid to be made available Open Access, so that it is freely accessible to all. The article was published online in the prestigious journal ANNALS of Neurology on the 31 December 2012.

Paper reference:

McGown, A. et al. Early Interneuron Dysfunction in ALS: Insights from a mutant sod1 Zebrafish Model. ANNALS of Neurology 2012 DOI: 10.1002/ana.23780
http://onlinelibrary.wiley.com/doi/10.1002/ana.23780/abstract

Mastering Pac-Man

Growing up in a seaside town in the 1980s led to me spending a lot of my “formative years” in the local amusement arcades, playing iconic video games like Space Invaders (at which I was average) and Asteroids  (I was the kid to beat!). One game I never got the hang of was Pac Man, where you had to guide a munching yellow ball around a maze eating up lots of dots. I even bought a book ‘Mastering Pac Man’ which didn’t help much – I was just plain rubbish!

Dr Brian Dickie and Pac Man

Talking about rubbish, fast forward 30 years and Prof Anne Simondsen from The University of Oslo is on the platform in Chicago describing the ways in which neurones deal with their cellular rubbish, such as poorly manufactured or damaged proteins.

Much as the dots are gobbled up by Pac Man, so cells employ a couple of basic ways to dispose of and recycle damaged proteins. One is a process called proteasomal degradation, which has been discussed on this website before by one of our guest bloggers, Prof John Mayer. However, some proteins, especially protein aggregates, are a bit too large and tough to be broken down by the proteasome, but they can be degraded by another process called autophagy – which can literally be translated as ‘self-eating’.

Prof Simondsen explained how aggregated proteins and even larger cellular structures can be packaged up for destruction by being encapsulated within membranes, forming structures called autophagosomes. These in turn seek out a structure called the lysosome, which contains digestive enzymes which break down the protein rubbish and keep the cell ‘spick and span’.

Using studies involving fruit fly models (a favourite model for cell biologists) she showed that the autophagy process declines with age, which is unfortunate because protein damage tends to increase with age. It was therefore no surprise that the level of autophagy in the brain was directly associated with the accumulation of protein ‘inclusions’, the health of neurones and the life expectancy of the flies.

Much of the evidence supporting a role for autophagy in neurodegeneration comes from the field of Huntington’s disease. Prof Simondsen explained how a particular mutated protein – called huntingdin – accumulates in the dying neurones and is a classic pathological hallmark of the disease. She demonstrated that autophagy normally plays a central role in the disposal of damaged huntingdin, but the system simply cannot cope with the amount of damaged protein, which starts to accumulate and literally ‘gunge up’ the cell. However, by stimulating the manufacture of additional autophagy machinery, the amount of aggregated protein can be reduced, helping to protect the neurones. This has so far only been shown in simple lab models of Huntington’s disease, but the encouraging results mean that further development to try and develop a treatment is underway.

So, could a similar approach be useful in other neurodegenerative diseases such as MND? Certainly, for some types of MND, such as those linked to a rare gene mutation called CHMP2B, it appears that a component of the autophagic machinery is impaired, which leads to protein build-up in cell models.

Dr Eiichi Tokada (Umea University, Sweden) provided further evidence that autophagy plays a role in disease progression in the SOD1 form of MND. By switching off a crucial component of the autophagy machinery, the disease progression in SOD1 mice was accelerated and their lifespan shortened. In addition, when he examined the spinal cords of the mice, he saw an increased presence of the characteristic SOD1 protein aggregates – a pathological hallmark of the disease.

Dr Faisal Fecto (Northwestern University, USA) provided evidence from a third genetic cause of MND. He showed how mutations in the Ubiquilin 2 gene disrupt the autophagy pathway by stopping the autophagosomes from linking up with the lysosomes.

So, it certainly seems that autophagy has a role to play in some of the rarer familial froms of MND. It remains to be seen to what extent it is involved in MND more generally, but it may mean that the potential treatment strategies being developed for Huntington’s disease may offer future opportunities for MND as well.

Our International Symposium website news stories:

International Symposium closes in Chicago

International Symposium focuses on clinical trials

International Symposium focuses on carer and family support

International Symposium begins in Chicago

Researchers unite at our International Symposium on MND

After you’ve finished reading the symposium articles that interest you, we’d be grateful if you could spare a few minutes to fill in our short online survey on our symposium reporting. Your comments really are useful and allow us to continually improve our symposium reporting. surveymonkey.com/s/alssymp

A prize-winning story worth repeating

Many congratulations to Rosa Rademakers from Mayo Clinic Florida USA, winner of this year’s Paulo Gontijo Young Investigator award. She won the award for her work on co-discovering the gene defect in C9orf72.

As part of her prize (in addition to a medal and a cheque to continue her work) she gave an overview of the research at the opening session of the 23^rd International Symposium on ALS/MND. The story was one of looking in some unusual places as well as all the obvious places to locate a gene defect had been thoroughly searched by researchers around the world. Dr Mariely DeJesus Hernandez in Dr Rademakers lab spotted something odd about the way the C9orf72 gene was inherited from the respective parents of someone with MND. She should’ve seen the copy from the mother and the copy from the father, but using their usual laboratory experiment, a copy of the gene from one of the parents wasn’t found.

One explanation for this unusual finding was that there was a ‘repeat’ sequence – that the experiment she’d run wasn’t set up to find. So, thanks to all the previous reports in the literature, Dr Rademakers and colleagues tried a lab experiment that other people had used to detect repeat sequences in other (ie non-MND) diseases. Use of this new lab experiment led to them identifying the presence of a long repeat in people with MND but not in unaffected people.

After a brief history of the discovery of this important gene defect, Dr Rademakers went on to give an overview of research around the world. It was interesting to see that this has worldwide significance. She showed a graph representing the percentage of cases of people with a family history of MND where C9orf72 had been discovered. The bottom line was that C9orf72 repeats are found in 34% of people who had MND with a family history of the disease and in 26% of people who had FTD with a family history of the disease.

But although much has been achieved in identifying this gene defect and the colossal amount of work worldwide since its discovery, in her final slide, Dr Rademakers reminded us that there’s much still to be done. For every concluding comment there was a list of two or three questions that the new information provoked.

This talk was an excellent starting point for a topic that was and will be repeated many times (pun intended) through the International Symposium.

Our International Symposium website news stories:

International Symposium closes in Chicago

International Symposium focuses on clinical trials

International Symposium focuses on carer and family support

International Symposium begins in Chicago

Researchers unite at our International Symposium on MND

After you’ve finished reading the symposium articles that interest you, we’d be grateful if you could spare a few minutes to fill in our short online survey on our symposium reporting. Your comments really are useful and allow us to continually improve our symposium reporting. surveymonkey.com/s/alssymp

University of Bath research shows how angiogenin affects motor neurone survival

Published in the prestigious journal Nature Communications and funded by the Wellcome Trust, University of Bath researchers looked closely at the structure of 11 mutated angiogenin proteins, and how changes in their structure influenced activity, function and survival in motor neurones.

Dr Brian Dickie, Director of Research at the Association, said “The researchers at the University of Bath have skilfully combined aspects of biology, chemistry and physics to answer some fundamental questions on how angiogenin can damage motor neurones. It not only advances our understanding of the disease, but may also give rise to new ideas on treatment development.”

Mutated forms of the gene editing protein, angiogenin, are known to cause MND in some families affected by the inherited form of the disease. This is due to a mutation in the normal angiogenin gene (a mistake in the instructions for making the protein) that leads to the production of a faulty/mutated angiogenin protein.

This new research expands on our knowledge of angiogenin, as back in June this year angiogenin, was identified by Irish researchers as playing a vital role in motor neurones with the effects tested in a mouse model of MND.

Creating images with x-rays

University of Bath researchers used a technique known as x-ray crystallography (a method that uses x-ray light to produce highly detailed images of the structure of a protein). Using this technique they were able to view the structure of angiogenin, and 11 angiogenin MND mutants, in exquisite detail. Specifically the researchers looked at the active site (where all the action happens) for changes that may affect the function and activity of the mutated proteins.

These x-ray crystallography images show stunning structural differences in the angiogenin MND mutant proteins, particularly around the active site, and the researchers found that these structural changes caused the mutated angiogenin proteins to have varying levels of activity.

Dr Vasanta Subramanian, who was involved in the research, said: “We hope that the scientific community can use this knowledge to help design more drugs that will bind selectively to the defective protein to protect the body from its damaging effects.”

An MND Association funded study has also used this technique of x-ray crystallography to look at the structure of mutant SOD1 proteins, which are another cause of inherited MND. Under the leadership of Prof Samar Hasnain, Dr Neil Kershaw is also one step ahead and is examining potential drugs to ‘mend’ these abnormal SOD1 proteins. This shows how this type of research, investigating structure and activity of the mutated proteins, can influence the development of new drugs.

What happens to angiogenin in the cell?

As well as changes in the activity of the protein the structural changes cause the angiogenin MND mutants to lose their function, leaving a protein sitting around in the cell, unable to do anything! The researchers found that it is essential that angiogenin is transported from the cytoplasm to the nucleus of the cell in order for the protein to function properly. This is an important step during the process in which proteins are made, if this does not occur then the protein will not function properly. In the angiogenin MND mutant proteins this does not occur correctly with either none, or very little, angiogenin being transported to the cell nucleus. This means that the angiogenin MND mutant proteins do not function properly compared to the normal angiogenin protein.

Stress Granules

The University of Bath researchers also found that the angiogenin MND mutant proteins prevented the motor neurones from producing ‘stress granules’. These are little packages that appear when a neurone is under stress, such as when low oxygen levels occur. When this stress factor occurs they release their contents to protect the motor neurone from damage.

These stress granules are the motor neurones natural defence mechanism. Without these granules the cell’s natural defences are lost and this influences the survival of the cell.

In Summary

This research gives us an important insight into how MND mutations affect the normal structure and function of angiogenin in motor neurones in some inherited forms of the disease. This not only advances our understanding of the disease, but may give rise to new ideas on treatment development in the future.

University of Bath press release

Journal article

Research we fund

EPHA4 gene influences survival in MND

An international research group spanning seven countries and including 23 researchers has identified a gene that modifies survival in MND. The gene, called EPHA4 was identified through a zebrafish genetic screening project and verified in rodents and humans with MND. The study was led by Prof Wim Robberecht, who has previously been funded by the MND Association and who is the Chair of our International Symposium on ALS/MND. The findings were published in the prestigious journal Nature Medicine this week.

What did the research group find?

By screening zebrafish for genetic factors that can modify the progression of MND, the research group identified EPHA4. By stopping, or slowing down the activity of EPHA4, they identified that MND zebrafish can be rescued and rodents (mice and rats) can live longer.

They also identified that MND vulnerable motor neurones have a higher level of EPHA4 than those at a lesser risk of developing the disease. This also means that a low level of EPHA4 confers to a lower risk of MND.

The research group then looked to humans to see if anybody with MND had mistakes in the EPHA4 gene that resulted in a change in survival. The group found two people with MND with genetic differences in the EPHA4 gene. These two people lived with MND for an exceptionally long time. As these genetic differences result in a lower level of EPHA4, this suggests that EPHA4 could be a valid therapeutic target for MND.

What does EPHA4 do?

Ephrin type-A receptor 4 (EPHA4 for short), plays a vital role in the development of our nervous system, in maintaining the shape of the neurone and in preventing regeneration after injury.

As our motor neurones grow, the projecting length of the neurone (the axon) needs to be guided to grow toward the right areas to connect to its respective muscle. To do this, a complex ephrin negative signalling system is used to guide the growing neurone in the right direction.

In real life terms, this signalling pathway can be thought of as somebody blindfolded, navigating a traffic cone maze. This person (the neurone) doesn’t want to move into an area cornered off by traffic cones (the corresponding ephrin signal). As the neurone can’t see where it’s going, it feels its way around using Eph receptors like EPHA4. The neurone moves away from ephrin signals as it ‘feels’ them. This eventually leads to the neurone reaching its target destination (the muscle).

To physically make the neurone move, when an ephrin signal connects to the ephrin receptor, the inner workings of the neurone are called into action. To find out more information on this, please read our previous article about Profilin1, which was found to be a cause of MND last month.

As EPHA4 plays an integral role in stopping neurone growth, it isn’t surprising that it also plays a role in stopping regeneration of neurones after injury. For example, in mice that have spinal cord injury (this was a study unrelated to MND), by genetically stopping the EPHA4 signal, new axon growth can be seen. With mice that have spinal cord injury that have a normal level of EPHA4 signal, growth cannot be seen.

The above non-MND study, along with the current EPHA4 finding further suggests that a lower level of EPHA4 can result in a longer survival because of its inability to perform its usual function to its usual extent. This incompetance seemingly allows the protection of neurones from degenerating at it’s normal pace.

As an additional note, you may be thinking that it seems counterintuitive that humans have additional signals that stop processes like regeneration when less complex animals like frogs still have this ability. Unfortunately, it’s a bi-product of mammalian evolution that continues to baffle scientists!

What does this mean for people with MND?

This finding unfortunately does not mean that a new genetic test will become available for EPHA4.

This discovery does however, offer another target for research institutes to look into and develop therapies that could slow down disease progression by lowering the amount of EPHA4.

Although this research is likely to take years to develop toward a clinical trial* in humans, it’s promising to see yet another exciting genetic advance that could have an impact on finding a better treatment for MND in the future.

*It’s important to point out that therapeutically lowering the EPHA4 signal in humans would not necessarily mean that neurones could regenerate as seen in the zebrafish. Many different signals other than EPHA4 prevent a human motor neurone from regenerating and from finding its target muscle. However, finding a way to lower the EPHA4 signal may still slow down progression, as seen in mice in this study.

What does this mean for the future of MND research?

Further studies are needed to verify and expand on these exciting results.

This finding means that researchers can explore this pathway in more detail as it, in conjunction with the recent Profilin 1 finding, suggests that this guidance/growth system of motor neurones may play an important role in the development of MND.

References:
Van Hoecke et al. Nature Medicine 2012 doi:10.1038/nm.2901

Munro et al. PLoS One 2012 10.1371/journal.pone.0037635

Our blog article on Profilin1

Profilin 1 identified as a cause of inherited MND

An MND causing gene called profilin 1 has been identified as the cause of about two percent of cases of inherited MND. This finding provides new insights into the causes of MND and suggests a potential role of the cellular scaffolding in MND. The finding was published in the 15 July edition of the prestigious journal Nature. This international collaborative study was led by Dr John Landers, at the University of Massachusetts Medical School, USA.

Using cutting-edge genetic technology, Dr Landers and colleagues first identified genetic mistakes in the profilin 1 gene in two families with inherited MND. To verify these findings, they went on to identify five additional families that also have mistakes in the profilin 1 gene. They did this by examining the genetic spelling of this gene in 272 further people with inherited MND (with no known genetic cause). This means that the genetic mistake could account for approximately two percent of cases of inherited MND.

Four different genetic mistakes were identified in the profilin 1 gene in the seven identified MND families. Three of these genetic mistakes were not found in any healthy controls, which mean that these mistakes are most likely a direct cause of MND. The forth genetic mistake was identified in a small number of healthy control samples, which could mean that this mistake could be a less significant cause of MND.

What does profilin do?

Profilin plays a vital role in maintaining and shaping the cells scaffolding – the cytoskeleton.

The cytoskeleton can be thought of as being made up of stacks of Lego bricks, called filaments. To maintain the shape of the cell, these bricks push against the cell membrane. To stretch and move the cell, more bricks (called actin) are added to the outermost end of the filament, which forces the membrane to extend. Toward the innermost end of the filament, the actin units separate, similar to pulling off individual bricks from the bottom of a stack, where they’re then collected and attached to profilin. Profilin then recharges and recycles the actin units, so that they’re ready to be added to the top of the filament again.

What did the research group find?

Through this study, the research group identified that the ability of profilin to attach to actin is affected by the genetic mistakes, making it ‘clumsy’. They also identified that the mistakes affect the ability of the cells to grow, which could be an attributing factor to how these mistakes can cause MND.

In this study, the researchers also confirmed that profilin is normally found throughout the ‘factory floor’ of the cell, the cytoplasm. However, when profilin is faulty the research group identified that it often assembles into clumps of protein marked for destruction – a hallmark of MND.

Interestingly, they also identified that when profilin is faulty, TDP-43 also clumps together. This suggests that faulty profilin may also cause MND through its effect on TDP-43. It’s also worth noting that when TDP-43 is faulty, profilin is not found within the clumps of faulty TDP-43 suggesting that profilin has an effect on TDP-43 and not vice-versa.

What does this mean for people with MND?

Profilin 1 is the twelfth MND causing gene to be identified in MND, which means that we are one step closer to knowing all of the genetic causes of MND. Learning more about how genetic mistakes can cause the rare inherited form of MND (5-10% of cases)  helps us to learn more about all forms of MND as the more common sporadic form is clinically indistinguishable to the inherited form.

As this genetic mistake is thought to only be attributed to a small number of families with MND, it is currently unknown if a genetic test will be developed for inherited MND. If you have inherited MND and want to find out more information about genetic testing, please speak with your doctor or neurologist.

What does this mean for the future of MND research?

These findings will need to be verified in larger numbers in different populations to determine a more accurate figure for how many families are affected by mistakes in the profilin 1 gene. More work will also need to be done to determine how the cytoskeleton is affected in MND and whether it can provide any therapeutic targets to treat MND in the future.

In summary, problems with the cytoskeleton have long been thought to be involved with MND, but having a direct genetic cause of MND strongly associated with the cytoskeleton will most likely reignite this avenue of research in the coming years.

Reference:
Wu C-H et al. Mutations in the profilin 1 gene cause familial amyotrophic lateral sclerosis. Nature 2012 doi:10.1038/nature11280